The Cytoskeleton in Normal and in Trans- formed Cells

Research Program A
Cell Differentiation and Carcinogenesis
Division A0100
Cell Biology
Division Cell Biology (A0100 / A010)
Head: Prof. Dr. Werner W. Franke
14
The Cytoskeleton in Normal and in Transformed Cells: Molecular Characterization of
the Main Components, Functional Domain
Analysis and Tumor Diagnosis
Scientists:
Dr. Hans Heid
PD Dr. Harald Herrmann-Lerdon
Dr. Ilse Hofmann
Prof. Dr. Jürgen Kartenbeck
Dr. Sandra Kneissel (3/02-)
Dr. Lutz Langbein
Dr. Ulrich-Frank Pape (3/02-)
Wiebke Peitsch (3-12/02)
PD Dr. Marion Schmidt-Zachmann
W.W. Franke, H. Herrmann-Lerdon, I. Hofmann
L. Langbein
Graduate students (Ph.D./Diploma/M.D.):
Kemal Akat
Angelika Alonso (3/01-)
Judit Boda (10/01-)
Christine Dreger
Jens Eilbracht
Olga Freidekind (6/02-)
Bettina Hämmerling (12/01-) Cathleen Hanisch (12/02-)
Alexandra König
Ute Patzelt (6/02-)
Michaela Reichenzeller
Holger Schlüter (4/02-)
Jens Schumacher (2/02-)
Beate Straub
Susanne Voltmer (8/02-)
Technical Assistants
Jutta Arlt (85%; -05/02)
Peter Eichhorn
Ulrike Figge (75%)
Christine Grund (80%)
Uta Haselmann-Weiss (3/02-) Michaela Hergt
Astrid Hofmann
Andreas Hunziker (-5/01)
Cäcilia Kuhn
Edeltraut Noffz (50%)
Jutta Osterholt
Silke Prätzel
Tanja Schlechter (2/02-)
Heiderose Schumacher (50%)
Tatjana Wedig
Stefanie Winter-Simanowski (75%)
Ralf Zimbelmann
The Division of Cell Biology investigates intracellular architectonic elements, both in the nucleus (karyoskeleton) and
the cytoplasm (cytoskeleton), and their interactions with
the nuclear envelope and cell junctions, respectively. The
constituting proteins, their interactions and functions are
characterized by use of biochemical methods, recombinant DNA technology and morphological analyses, including electron microscopy and immunolocalization techniques, and their patterns of synthesis in normal and malignant cells and tissues are determined. To elucidate possible functions of these proteins and structures gene
transfer and expression techniques are applied, including
stably transfected cell lines as well as transgenic and
gene-targeted mice, to solve principal problems of cell biology that are central to the understanding of cell and tissue differentiation, carcinogenesis and metastasis. The reagents generated, in particular monoclonal antibodies developed in the course of these studies, are also examined
for their diagnostic value in the detection and classification
of tumors.
In cooperation with: Prof. Dr. Ueli Aebi, Drs. Sergei V. Strelkov,
Peter Burkhard, Biozentrum, Basel, Switzerland; Drs. M. Amagai,
T. Hashimoto, Keio University School of Medicine, Tokyo, Japan;
Prof. Dr. H.-H. Arnold, University of Braunschweig; Prof. Dr.
Helene Baribault, La Jolla Cancer Research Foundation, USA;
Profs. Drs. Avri Ben Ze’ev, Benny Geiger, Weizmann Institute of
Science, Rehovot, Israel; Prof. Dr. Walter Birchmeier, MDC für
Molekulare Medizin, Berlin; Dr. B. Cribier, Dept. Dermatology,
Université de Strasbourg, France; Profs. Drs. Helmut Denk, Kurt
Zatloukal, Dept. Pathology, University of Graz, Austria; Prof. Dr.
Robert D. Goldman, Northwestern University Medical School,
Chicago, USA; Prof. Dr. Victor E. Gould, Department of Pathology, Rush-Presbyterian-St. Luke’s Medical Center, Chicago, USA;
Drs. M. Guerrin, M. Simon, Purpan School of Medicine, Toulouse
University, Toulouse, France; Prof. Dr. Walter Keller, Dr. Isabelle
Kaufmann, Biozentrum, University of Basel, Switzerland; Prof. Dr.
Peter Krammer, DKFZ; Dr. M. Krawczak, Inst. Medical Genetics,
University of Wales, Cardiff, U.K.; Dr. Hjalmar Kurzen; Dept. Dermatology, Heidelberg University Medical School; Prof. Dr. Irene
M. Leigh, Skin Tumor Lab., Royal School of Medicine, London,
U.K.; Prof. Dr. Peter Lichter, DKFZ; Prof. Dr. Thomas Magin,
Institut für Genetik, Universität Bonn; Prof. Dr. Jürgen Markl, Dr.
J. Robin Harris, Zoologisches Institut, Universität Mainz; Prof. Dr.
Ingrid Moll, Hautklinik und Poliklinik, UKE Hamburg; Prof. Dr.
Roland Moll, Dr. Ansgar Schmidt, Medizinisches Zentrum für
Pathologie, Universität Marburg; Drs. Ada and Don Olins,
Bowdoin College, Biology Department, Brunswick, USA; Prof. Dr.
Robert G. Roeder, New York, USA; Dr. Valentino Romano,
Laboratorio di Genetica Moleculare, Troina, Italy; Prof. Dr. Dennis
Roop, Dept. Molecular and Cellular Biology and Dermatology,
Baylor College, Houston, USA; Drs. Jürgen Schweizer, Michael A.
Rogers, Hermelita Winter, DKFZ; Drs. Y. Shimomura, N. Aoki,
Niigata University School of Medicine, Dept. Dermatology,
Niigata, Japan; Dr. Herbert Spring, DKFZ; Prof. Dr. Annegret
Starzinski-Powitz, Humangenetik, Universität Frankfurt; Dr. Ole
Swensson, Hautklinik, Universität Kiel; Drs. A. Taieb, V.
Chapalain, Unité de Dermatologie, Bordeaux, France; Prof. Dr.
Jean-Paul Thiery, CNRS, Laboratoire de Physiopathologie du
Développement, Paris, France; Dr. Eishin Yaoita, Institute of
Nephrology, Niigata University, Japan.
Cellular architecture and the interaction of individual cells
with neighboring cells or extracellular matrix components
is mediated by cell type-specific protein complexes termed
the “cytoskeleton”. The mechanisms of the cell type- and
differentiation-specific synthesis of sets of cytoskeletal
proteins, their assembly and coordinated integration into
various structures differ considerably in different kinds of
cells and tissues, both in the normal and the transformed
situation. In order to understand a specific developmental
process and the maintenance of the differentiated state it
is of fundamental importance to elucidate the contribution
and localization of individual architectonic molecules, their
modifications, and the nature of their functional domains
that contribute to the specific cell structure and the functions. In particular, we investigate the specific expression
in neoplastic cells and the importance of biological tools
e.g. specific antibodies for clinical tumor diagnosis.
DKFZ 2003: Research Report 2001/2002
Research Program A
Cell Differentiation and Carcinogenesis
I. Assembly, structure and function of
intermediate filaments
As a model protein to study general principles of IF assembly we have chosen the type III IF protein vimentin
and shown it to polymerize in three distinct phases [20].
However, also cytokeratins, desmin and neurofilament
proteins were shown to follow the same assembly schedule starting from so-called “unit-length filaments” [21].
Most notably, we obtained the first atomic structure information of IF proteins crystallizing the IF consensus sequence of vimentin [54, 55]. The assembly of certain IF
proteins is temperature-dependent as has been shown for
vimentin from various species [50].
This extraordinary degree of temperature-sensitivity of
vimentin has been utilized in a novel context. The existence of a hitherto more or less hypothetical space between the chromosomes of the interphase nucleus (the interchromosomal domain compartment) was visualized and
explored by directly employing a “nuclear space shuttle”,
namely Xenopus vimentin carrying a nuclear localization
signal introduced by recombinant DNA techniques [9, 46].
In stably transfected cultured human cells, the transgene
product is deposited in nuclear aggregates at 37°C. However, upon lowering the growth temperature to 28°C, filaments start to grow out of these aggregates connecting
them within a few hours. The “network” of filaments formed
this way is found exclusively outside of chromosomal territories colocalizing with various nuclear entities such as
coiled bodies, PML bodies or newly transcribed RNA.
The cytoskeleton is also intimately engaged in drastic
changes and transitions of nuclear architecture, as observed during the differentiation of certain blood stem
cells. Thus, during the generation of granulocytic cells
from undifferentiated precursors, in the course of which
nuclear shape and probably function changes by complex
lobulations and evaginations, the vimentin in the cytoplasm and the lamin system in the nucleus as well as the
lamin receptors are fundamentally affected [39]. Similarly,
also intranuclear lamin filaments and inner nuclear membrane proteins such as the lamin B receptor, LBR, are intimately engaged in the architectural organization of the
nucleus being reflected by the evolutionarily conserved
molecular structure of these IF proteins and their central
importance during oogenesis and embryogenesis [23, 51].
The integration of IFs into the cellular structure, i.e. the cytoskeleton, involves a number of associated proteins one
of which is plectin. Its central role in cellular structure and
physiology has been shown in the context of the apoptotic
reorganization process and during viral infection [18].
II. Keratin-intermediate filaments and associated
proteins - Expression and function in special
epidermal compartments
Intermediate filament (IF) proteins represent - like the
other filament-forming proteins - a multigene family with
more than 50 members, which are expressed in cell typeand differentiation-specific patterns, constitute scaffolds
with essential structural functions in the cytoplasm (e.g.,
keratins in epithelial cells, vimentin in mesenchymal cells,
desmin in muscle cells, glial and neurofilament proteins in
Division A0100
Cell Biology
glial and neuronal cells, respectively). The (“soft” or cyto)epithelial keratins (Ks) and the (“hard”) hair keratins (HKs)
are subgroups of the large keratin IF-protein multigene
family. Whereas the epithelial keratins are cytoskeletal
components of keratinocytes and other epithelial cells, the
HKs are found in specific keratinocytes, trichocytes, of the
hair and nail forming compartment.
Extending our previous work of the expression of the type I
HKs (hHa), which consists of 9 hHa (hHa1-hHa8; including
the two rather related proteins hHa3-I, hHa3-II) a major
aim was the completion of the HK gene expression and
therefore we investigated the type II HKs (hHb). Previously
only four members of the human type II HKs polypeptides
were known at biochemical (protein) level. Now we have
shown that there exist 6 hHb (hHb1-6) [34]. Large gene
expression studies by using in situ hybridization and immunohistochemistry with specific antibodies produced in
our laboratory have demonstrated that the HKs - comparable to the epithelial keratins - show patterns of sequentially and differentiation-dependent expression within the
hair forming compartment, the hair follicle: The expression
of some members (hHb5 + hHa5) begin in the upper matrix cells, a region formerly assumed to be free of keratins.
The synthesis of other HKs follows in the lower/mid cortex
(hHb1, hHb3, hHb6 + hHa1, hHa3-I, hHa3-II) and ends in
the upper cortex (hHa4). Moreover, the HKs hHa2 and
Hhb2 are exclusively synthesized in the hair cuticle. In
contrast, hHb4, although a true hair keratin, is not expressed in the hair follicle but in the filiform papillae of
tongue. Based on these results and aware that all of the
HK genes are known we now established the catalog of
human type-I and type-II HKs [34].
One of the type I HKs, the pseudogene ΨhHaA, is transcribed in humans but do not exist as a functional protein
because of a premature stop codon. It is, however, clearly
detectable as a functional gene in chimpanzee hairs and during evolution only about 240,000 years ago - homozygoteously mutated and lost in the hominide lineage [60].
Furthermore, the work was expanded to study aspects of
the gene regulation of HKs and described the role of the
transcription factor HoxC13 in this respect [28].
First experiments were done on the filament formation of
hair follicle keratins in vitro. Both forms of keratins, cortex
and companion layer keratins, form heteropolymeric complexes in 5M urea. Whereas K6hf and K17 form also filaments in low salt buffers the HKs tested need physiological salt conditions. Moreover, point mutated HKs were not
able to form long filaments but only “unit length” filaments
[25].
Recently we have found K6hf, that is specifically expressed in the companion layer of the hair follicle and
have - in contrast to earlier studies, who named this structure “innermost layer of outer root sheath” - demonstrated
that this compartment is an own, single-layered differentiation sui generis and not part of the outer root sheath. Interestingly, CK6hf seems to be involved in a hair disease
named “loose anagen” [6].
During our studies we also detected a new epithelial keratin, K6irs1. This keratin is restricted to the inner root
DKFZ 2003: Research Report 2001/2002
15
Research Program A
Cell Differentiation and Carcinogenesis
16
Division A0100
Cell Biology
sheath (IRS) of the hair follicle, i.e. all three compartments, the Henle-, Huxley- and IRS cuticle layer. This protein is also found in specialized cells of the Huxley layer,
named Flügelzellen (“winged cells”). These Huxley cells
pass the Henle layer and abut on companion layer by numerous desmosomes and insert flexible elements in the
otherwise rigid Henle layer [36]. Furthermore, we also
found three new keratins clearly restricted to specific compartments within the IRS, with K6irs2, K6irs3 sequentially
expressed in the IRS-cuticle and K6irs4 in the IRS Huxley
layer [Langbein et al., 2003, in press].
proteins, which are valuable in the immunohistochemical
differentiation and identification of various cell types in
normal development and in tumor diagnoses (several ms.
submitted). This panel of antibodies to junction type-specific components of both the plaque and the extracellular
segments has also allowed to detect and define novel
kinds of adhering junctions which cannot be subsumed
under the hitherto existing categories of “adherens” and
desmosomal junctions.
The work on HKs are also the basis for the characterization and identification of tumors originated from the hair
forming compartment (see also chapter V of this report).
The occurrence of extended tight junction (TJ) structures,
including zonulae occludentes (ZO) in stratified cornified
and non-cornified epithelia was long and controversially
discussed as their existence was only accepted in polar
(non-stratified) epithelia. Therefore, we have systematically examined the localization of TJ proteins in diverse
stratified epithelial tissues of various species as well as
cell culture lines derived from stratified epithelia, by electron microscopy as well as by immunocytochemistry at
both the light and the electron microscopic level, using antibodies to TJ proteins such as occludin, claudins 1 and 4,
protein ZO-1, cingulin and symplekin [4, 35]. Unexpectedly, besides “classical” TJ-protein positive structures we
have found a diversity of TJ-related structures of variable
sizes especially in the upper layers of such epithelia:
“lamellated TJs”, coniunctiones laminosae (junctional regions showing relatively broad, ribbon-like membrane contacts which in cross-section often appear pentalaminar
with an electron-dense middle lamella), “sandwich junctions”; iuncturae structae (characterized by a 10-30 nm
dense lamina interposed between the two membranes),
and “cross-bridged cell walls”; parietes transtillati (with a
intermembrane distance which is rather regularly bridged
by short rod-like elements). These structures were often
intermingled with each other and mostly in close vicinity to
desmosomes. In part, they resemble structures formerly
described as other types (e.g. gap-junctions) of cell-cell
junctions [35]. Although often existing as widely extended
structures they were obviously overlooked for a long time.
These TJs and TJ-related structures are now considered
as being part of the barrier system in stratified epithelia.
Besides the expression studies of HKs we also investigated that of keratin-associated proteins (KAPs), proteins
which cross-link keratin filaments to built of the rigid structure of the hair fibre. They also exist as multigene families
which include among others the high/ultrahigh sulfur KAPs
[47, 52] and glycine/tyrosine KAPs [48] with one race specific polymorphism [53].
III. Desmosomes - intercellular connecting
structures and anchoring sites of the
cytoskeleton
Our studies on the molecular composition and functions of
intercellular junctions, particularly those of the “adherens”
group, have focussed on the clarification of the complexity,
localization and functions of proteins of the so-called “armrepeat” family, known to be important cytoplasmic components of junctional plaques and of the transmembrane proteins, the desmosomal cadherins, the desmogleins and
desmocollins.
To examine the function of various transmembrane proteins localized in adhering junctions the genes of desmoglein 2 (DSG2), an ubiquitously expressed desmosomal
protein, and M-cadherin, a classical cadherin found in
muscle and brain, were inactivated by homologous recombination in embryonal stem cells for the generation of
knock out mice. Germline inactivation of DSG2 lead to embryonic lethality whereas M-cadherin deficient mice were
viable and fertile and showed no gross developmental defects [11, 27].
In recent years we noticed that several components of
junctional plaques are also found in karyoplasmic, interchromatinal granules, even in cells devoid of any junctions. By biochemical methods the nuclear form of the
demosomal plakophilin 2 has been detected in specific
complexes with the largest subunit of RNA polymerase III
[37]. The nuclear and cytoplasmic form of the protein
symplekin, which is also located at tight junctions, is associated with proteins involved in mRNA biogenesis, notably
3’-end processing in the nucleus and regulated polyadenylation in the cytoplasm [26]. Interestingly, both characterized nuclear form of junctional plaque proteins have
in common that they are involved in processes of transcription and processing of pre-mRNA.
Furthermore, we have developed several types of antibodies specific for individual desmosomal cadherins, i.e. desmogleins 1-3 and desmocollins 1-3, and other junctional
IV. Tight junctions and tight junction-related
structures in stratified epithelia
These studies were extended to investigate the situation in
carcinomas derived from such epithelia (see part V.).
V. Cytoskeletal proteins as markers of cellular
differentiation in tumor diagnostics
Antibodies against cytoskeletal elements e.g. keratins
were used to characterize the type and the differentiation
stage of normal and in particular neoplastic cells for a long
time. After characterization of the specific expression patterns of hair keratins and hair follicle- specific keratins
(e.g. K6hf, K6irs1-4) investigations were started to further
characterize carcinomas. Using antibodies against HKs
we could clearly identify e.g. pilomatricoma as being originated from the follicular compartment of the skin [8]. Furthermore, we investigated a panel of other carcinoma including trichoepithelioma and basal cell carcinoma [32]
and are also looking for ektopic, but characteristic, expres-
DKFZ 2003: Research Report 2001/2002
Research Program A
Cell Differentiation and Carcinogenesis
sion of such keratins in other epithelial neoplasias possibly
useful for diagnosis.
In order to continue our studies on tight junctions and tight
junction-related structures as well as their molecular components we started investigations about the occurrence
and composition of such structures in carcinomas e.g.
squamous cell carcinomas of various origin. Here we
could show for the first time, that there exist compartments
within one tumor, in particular in the high differentiated areas, which are “encapsulated” by tight junctional structures. Therefore an functional barrier between compartments of one tumor can be assumed. This might be important to characterize the tumor itself and, furthermore, to be
important for the penetration of e.g. cytotoxic substances
applied during therapy of the tumors [Langbein at al.,
2003, in press].
Karyoskeletal Elements, Intranuclear
Architecture and Topogenesis of
Nuclear/Nucleolar Proteins
M.S. Schmidt-Zachmann
In cooperation with Michael Stöhr, Dr. Martina Schnölzer, PD Dr.
Hanswalter Zentgraf, Dr. Herbert Spring (DKFZ); Prof. Dr. Joseph
Gall, Carnegie Institution, Baltimore, USA; Prof. Dr. Angela
Krämer, University of Geneva, Switzerland
The compartmentalization of the eukaryotic cell and its
functions are based on the specific topogenesis of cellular
proteins. While in the past decade important molecular
principles concerning the nucleocytoplasmic distribution of
proteins have been elucidated, such as the presence of a
short, basic signal sequence, termed nuclear localization
signal (NLS) in many proteins, it remains to be determined
to what extent and how distinct amino acid sequence motifs govern the targeting of proteins to precise subnuclear
structures.
The nucleus contains, in addition to the chromosomal
structures, the nuclear envelope and the lamina, a number
of distinct karyoplasmic structures and bodies of which the
nucleolus is by far the most extensively studied. The
nucleolus which represents an accumulation of rDNA and
its transcription products together with a characteristic set
of proteins, is a complex nuclear substructure in which key
steps of ribosome biogenesis as well as assembly of other
kind of ribonucleoprotein particles take place. Whereas a
number of additional karyoplasmic structures (e.g. nuclear
speckles, PML-bodies, and Cajal bodies) have been characterized over the past decade, taking advantage of specific “marker” proteins, the nature and functions of many
others are still unknown.
Our studies are aimed at the identification and biochemical characterization of various nuclear bodies, fibrils or
other structural subcomponents, with a special emphasis
on the nucleolus.
Division A0100
Cell Biology
teins which are involved in maintaining or even contributing to the nucleolar structural integrity, i.e. proteins of the
“nucleolar skeleton”.
Recently, we succeeded in the identification of a novel
type of karyoskeletal protein and molecular marker for a
specific nucleolar substructure, i.e. a relatively thin cortical
layer forming a cage-like perinucleolar structure and consisting of a meshwork of patches and filaments that dissociates upon reduction of divalent cations. Whereas this
protein- termed NO145 - can be classified as a bona fide
karyoskeletal protein (resistant to high-salt/detergent extraction) it is also sensitive to regulated proteolysis. Remarkably, protein NO145 (originally identified in Xenopus
oocytes) is present throughout all stages of oogenesis but
is rapidly degraded during meiotic maturation, i.e. egg formation [30, 31]. The mechanism responsible for this phenomenon, which distinguishes NO145 from all other nucleolar proteins known so far, is still unknown and requires
further investigations. Furthermore, partly homologous and
structurally or functionally related proteins in distant species should be identified in the near future.
As outlined before, we are also interested in the mechanisms regulating the specific accumulation of proteins in
distinct intranuclear structures. Previous studies on the intranuclear topogenesis of nucleolar proteins (e.g. [Zirwes
et al., Mol. Biol. Cell 8 (1997) 231-248] and references
therein) had revealed that nucleolar accumulation of proteins is a two step process: (i) active transport into the cell
nucleus via a functional nuclear localization signal (NLS),
and (ii) accumulation in the nucleolus due to specific binding interactions between these proteins and other nucleolar components, particularly rDNA, rRNA and possibly
other protein constituents.
Recently, we have described the identification, cDNA cloning and immunodetection of a novel type of constitutive
nuclear protein which occurs in diverse vertebrate species,
from Xenopus to man, which becomes specifically enriched in nuclear speckles. Meanwhile the protein has
been identified as a subunit of the U2 snRNP-associated
splicing complex SF3b and is termed SF3b155 [SchmidtZachmann et al., Mol. Biol. Cell 9 (1998) 143-160]. We
have examined the topogenic properties of different protein domains of SF3b155 and have identified molecular segments that i) govern its nuclear import and ii) determine its
accumulation in nuclear speckles (mediated by the socalled “TP”-domain). This latter protein domain also contributes to the association of SF3b155 with the U2 snRNP
[10]. Our studies allows the conclusion that the mechanism of nuclear speckle localization is most likely a complex process with analogy to the pathways described for
localization of proteins in other nuclear substructures, e.g.
Cajal Bodies and nucleoli.
Over the years we have reported on the identification of
several constitutive nucleolar proteins (NO29, NO38,
NOH61), which have distinct functional roles during the
complex process of ribosome biogenesis. However, a major focus of these studies is the characterization of pro-
DKFZ 2003: Research Report 2001/2002
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Research Program A
Cell Differentiation and Carcinogenesis
Functions and Dynamics of Defined
Membrane Domains
J. Kartenbeck
18
In cooperation with: Prof. Dr. Angel Alonso, DKFZ; PD Dr. Ursula
Bantel-Schaal, DKFZ; Dr. Melanie Ott, DKFZ; Dr. Franz Bosch,
Dept. Otolaryngology, University of Heidelberg Medical School;
PD Dr. Nikolaus Gassler, Institute of Pathology, University of
Heidelberg; Prof. Dr. Ari Helenius, ETH Zürich, Switzerland; Prof.
Dr. Rudolf Leube, Institute of Anatomy, University of Mainz.
In three different projects we have used the determination
of expression and synthesis of cell-cell and cell-matrix
contact components for tumor diagnosis, differentiation
and the evaluation of prognostic factors.
In squamous cell carcinoma (SCC) we noticed with the onset of invasive growth the upregulation of the hemidesmosomal proteins Bullous pemphigoid antigen 1 (BPA1) and
α6β4 integrin. These proteins, normally localized at basal
layers of keratinocytes, extended to the proliferative zone,
and also in tumors which no longer formed hemidesmosomal structures. The redirected expression of these proteins to the entire surface of many tumor cell layers indicated a loss of polarity and was found to allow a prediction
on the potential of the invasive tumor cells to metastasize
[19].
In contrast to the above-mentioned situation the evaluation of reduced expressions of proteins engaged in the formation of intercellular contacts is generally taken as an indicator for malignancy and as a prognostic factor. Therefore, we analyzed head and neck squamous cell carcinomas (HNSCC) from 190 patients for the expression of the
desmosmal proteins, desmoplakin and desmoglein, and
for the adherens junction protein, E-cadherin. The expression patterns of these proteins from primary tumors were
compared with metastases therefrom. We saw a great
spectrum of tumor differentiation and variability of expression patterns. In most cases metastases revealed expression patterns as their primary tumors, and changed expression of the various proteins and structures behaved in
a parallel manner. Statistical evaluation revealed that a
strict correlation between expression pattern and metastatic tumor events was not possible. In the great majority
of cases junctional proteins of both intercellular contact
types (desmosome, adherens junctions) were still present,
although often with a decreased expression pattern [3].
As changes of the intestinal mucosal barrier play a role in
the pathogenesis of inflammatory bowel disease (IBD) we
evaluated potential dysregulation of intercellular junctional
proteins on tissues obtained from patients with Crohn’s
disease and ulcerative colitis. Marked downregulation of
these proteins was found in actively inflamed IBD, while in
inactively inflamed regions only the adherens junction proteins E-cadherin and α-catenin were affected. Expression
of desmosomal or tight junction associated proteins appeared almost unchanged. The results revealed that
downregulation of junctional molecules is apparently associated with the inflammatory process and does not represent a primary phenomenon [14].
Investigations on the internalization of two non-enveloped
viruses, the adeno-associated virus type 5 (AAV5) and
Division A0100
Cell Biology
simian virus 40 (SV40), in cultured cells revealed hitherto
not identified endocytotic pathways. AAV5, mainly internalized by coated vesicles, was seen to be routed to the
Golgi area where they could be detected within cisternae
of the trans-Golgi network and within vesicles associated
with dictyosomal stacks [2]. SV40 internalization took
place via caveolae and small caveolin-1 containing
vesicles. It then entered larger organelles with non-acidic
pH which did not contain marker for endosomes, lysosomes, endoplasmic reticulum (ER) or Golgi cisternae. Further transport to membranes of the smooth ER took place
in caveolin-free tubular vesicles. The results demonstrate
the existence of a two-step transport through an intermediate organelle (termed caveosome) bypassing endosomal
and Golgi compartments [42].
Special Cell Type-specific Cytoskeleton
Structures
H. Heid, I. Hofmann, W.W. Franke
In cooperation with: Prof. Dr. Ricardo Benavente, Biozentrum,
University of Würzburg; Prof. Dr. Helmut Denk, Prof. Dr. Kurt
Zatloukal, Department of Pathology, University of Graz, Austria;
Prof. Dr. Wolfgang Hagmann, Tumorbiochemistry, DKFZ; Prof. Dr.
Brigitte Jockusch, Cell Biology at the Institute of Zoology, University of Braunschweig; Prof. Dr. Thomas W. Keenan, Virginia Polytechnic Institute, Blacksburg, USA; Prof. Dr. Thomas Magin, Institute of Physiological Chemistry, University of Bonn Medical
School; Prof. Dr. Ingrid Moll, Prof. Dr. Wolfgang Schulze, Division
of Andrology and Department of Dermatology, University of Hamburg Medical School; Prof. Dr. Roland Moll, Department of Pathology, University of Marburg; Dr. Martin Volkmann, Medical Clinic,
University of Heidelberg; Prof. Dr. Jürgen Wehland, GBF,
Braunschweig.
In continuation of our former research on the molecular
composition of the calyx structure of sperm heads of
mammalia and the identification and characterization of
the sperm-specific basic proteins cylicin I, cylicin II and
calicin, as well as the “capping protein β3” [Hess et al., J.
Cell Biol. 122 (1993) 1043-1052; Hess et al., Exp. Cell
Res. 218 (1995) 174-182; von Bülow et al., Exp. Cell Res.
233 (1997) 216-224] we have identified two actin-related
proteins (Arps) termed Arp-T1 and Arp-T2 and the selenoprotein PHPGx among several other so far unknown proteins. Specific antibodies localized the two Arps within the
calyx structure [17]. These two proteins are obviously new
types of actin-related proteins with new functions not involved in the initiation process of new actin filament formation. The importance and the involvement of these specific
proteins within the cytoskeletal structure of sperm and the
regulation of their synthesis will be examined in more detail. Reports on sperm disorders (teratozoosperms; e.g.
so-called “round headers”) in cases when these proteins
are missing or wrongly arranged, give hints of the importance of such proteins.
Detection and characterization of novel proteins,
protein modifications and genes
In cooperation with other groups and through microsequencing of proteins and peptides several new proteins
were identified and characterized [31, 56, 58, 62].
DKFZ 2003: Research Report 2001/2002
Research Program A
Cell Differentiation and Carcinogenesis
Drebrin-containing structures in non-neuronal
cells and their function
We showed that the actin-binding protein drebrin, extending our previous report [Peitsch et al., Eur. J. Cell Biol. 78
(1999) 767-778], is actually a widespread protein present
in a wide range of non-neuronal cells and tissues occurring in distinct oligomeric complexes and in granules [40,
41]. These structures are often clustered at certain strategic sites in lamellipodia and filopodia suggestive of a functional involvement of drebrin in cell motility and shape
changes, notably formation of cell protrusions.
Publications (members of the Division in bold; * = external
co-author)
[1] Aleem, E.A., Flohr,T., Hunziker, A., Mayer, D., Bannasch, P.,
Thielmann, H.W. 2001. Detection and quantification of protein
phosphatase inhibitor-1 gene expression in total rat liver and isolated hepatocytes. Mol. Cell. Biochem. 217, 1-12.
[2] Bantel-Schaal, U., Hub, B., Kartenbeck, J. 2002. Endocytosis of adeno-associated virus type 5 leads to accumulation of virus particles in the Golgi compartment. J. Virol. 76, 2340-2349.
[3] *Bosch, F.X., *Schuhmann, A., Kartenbeck, J. 2001. On the
role of cell-cell adhesion in metastases formation in head and
neck cancer. In: Proceedings of the 2nd International Symposium
on “Metastases in Head and Neck Cancer”. B.H. Lippert and J.A.
Werner (eds.), Tectum Verlag.
[4] *Brandner, J.M., *Kief, S., Grund, C., *Rendl, M., *Houdek,
P., Kuhn, C., *Tschachler, E., Franke, W.W., *Moll, I. 2002. Organization and formation of the tight junction-system in human epidermis and cultured keratinocytes. Eur. J. Cell Biol. 81, 253-263.
[5] Cerdà, J., Grund, C., Franke, W.W., *Brand, M. 2002. Molecular characterization of calymmin, a novel notochord sheathassociated extracellular matrix protein in the zebrafish embryo.
Dev. Dyn. 224, 200-209.
[6] *Chapalain, V., Winter, H., Langbein, L., *LeRoy, J.M.,
*LabrÈze, C., *Nikolic, M., Schweizer, J., *TaÏeb, A. 2002. Is the
loose anagen hair syndrome a keratin disorder? A clinical and
molecular study. Arch. Dermatol. 138, 501-506.
[7] Cheng, H., Kartenbeck, J., Kabsch, K., Mao, X., Marqués,
M., Alonso, A. 2002. Stress kinase p38 mediates EGFR
transactivation by hyperosmolar concentrations of sorbitol. J. Cell.
Physiol. 192, 234-243.
[8] *Cribier, B., *Peltre, B., Langbein, L., Winter, H., Schweizer,
J., *Grosshans, E. 2001. Expression of type I hair keratins in follicular tumours. Br. J. Dermatol. 144, 977-982.
[9] Dreger, C.K., König, A.R., Spring, H., Lichter, P., Herrmann,
H. 2002. Investigation of nuclear architecture with a domain-presenting expression system. J. Struct. Biol. 140, 100-115.
Division A0100
Cell Biology
[14] *Gassler, N., *Rohr, C., *Schneider, A., Kartenbeck, J.,
*Bach, A., *Obermüller, N., *Otto, H.F., *Autschbach, F. 2001. Inflammatory bowel disease is associated with changes of
enterocytic junctions. Am. J. Physiol. Gastrointest. Liver Physiol.,
281, G216-G228.
[15] *Gassler, N., Schnölzer, M., *Rohr, C., *Helmke, B.,
Kartenbeck, J., *Grünewald, S., *Laage, R., *Schneider, A.,
*Kränzlin, B., *Bach, A., *Otto, H., *Autschbach, F. 2002. Expression of calnexin reflects Paneth cell differentiation and function.
Lab. Invest. 82, 1647-1659.
[16] *Grüttgen, A., Reichenzeller, M., *Jünger, M., *Schlien, S.,
*Affolter, A., *Bosch, F.X. 2001. Detailed gene expression analysis
but not microsatellite marker analysis of 9p21 reveals differential
defects in the INK4a gene locus in the majority of head and neck
cancers. J. Pathol. 194, 311-317.
[17] Heid, H.W., Figge, U., Winter, S., Kuhn, C., Zimbelmann,
R., Franke, W.W. 2002. Novel actin-related proteins Arp-T1 and
Arp-T2 as components of the cytoskeletal calyx of the mammalian sperm head. Exp. Cell Res. 279, 177-187.
[18] Henzler, T., Harmache, A., Herrmann, H., Spring, H.,
*Suzan, M., *Audoly, G., Panek, T., Bosch, V. 2001. Fully functional, naturally occurring and C-terminally truncated variant human immunodeficiency virus (HIV) Vif does not bind to HIV Gag
but influences intermediate filament structure. J. Gen. Virol. 82,
561-573.
[19] *Herold-Mende, C., Kartenbeck, J., *Tomakidi, P., *Bosch,
F.X. 2001. Metastatic growth of squamous cell carcinomas is correlated with upregulation and redistribution of hemidesmosomal
components. Cell Tissue Res. 306, 399-408.
[20] Herrmann, H., *Aebi, U. 2002. Stress-Fänger und Chromatin-Organisatoren: Hochdynamischen intrazellulären
Filamentsystemen auf der Spur. BIOspektrum 8, 597-601.
[21] Herrmann, H., Wedig, T., *Porter, R.M., *Lane, E.B., *Aebi,
U. 2002. Characterization of early assembly intermediates of recombinant human keratins. J. Struct. Biol. 137, 82-96.
[22] *Hofemeister, H., Kuhn, C., Franke, W.W., *Weber, K.,
*Stick, R. 2002. Conservation of the gene structure and membrane targeting signals of germ cell-specific lamin LIII in amphibians and fish. Eur. J. Cell Biol. 81, 51-60.
[23] *Hoffmann, K., Dreger, C.K., *Olins, A.L., *Olins, D.E.,
*Shultz, L.D., *Lucke, B., *Karl, H., *Kaps, R., *Müller, D., *Vayá,
A., *Aznar, J., *Ware, R.E., *Cruz, N.S., *Lindner, T.H.,
Herrmann, H., *Reis, A., *Sperling, K. 2002. Mutations in the
gene encoding the lamin B receptor produce an altered nuclear
morphology in granulocytes (Pelger-Huët anomaly). Nat. Genet.
31, 410-414.
[24] Hofmann, I. 2002. Interaktionen von Cytoskelett- und ZellZell-Verbindungsproteinen. Habilitation. Faculty of Biology, University of Heidelberg.
[10] Eilbracht, J., Schmidt-Zachmann, M. 2001. Identification of
a sequence element directing a protein to nuclear speckles. Proc.
Natl. Acad. Sci. USA 98, 3849-3854.
[25] Hofmann, I., Schnölzer, M., *Kaufmann, I., Franke, W.W.
2002. Symplekin, a constitutive protein of karyo- and cytoplasmic
particles involved in mRNA biogenesis in Xenopus laevis oocytes.
Mol. Biol. Cell 13, 1665-1676.
[11] Eshkind, L., Tian, Q., Schmidt, A., Franke, W.W.,
*Windoffer, R., *Leube, R.E. 2002. Loss of desmoglein 2 suggests essential functions for early embryonic development and
proliferation of embryonal stem cells. Eur. J. Cell Biol. 81, 592598.
[26] Hofmann, I., Winter, H., Mücke, N., Langowski, J.,
Schweizer, J. 2002. The in vitro assembly of hair follicle keratins:
comparison of cortex and companion layer keratins. Biol. Chem.
383, 1373-1381.
[12] *Ferrando-May, E., Cordes, V., *Biller-Ckovric, I., *Mirkovic,
J., *Görlich, D., *Nicotera, P. 2001. Caspases mediate
nucleoporin cleavage, but not early redistribution of nuclear transport factors and modulation of nuclear permeability in apoptosis.
Cell Death Differentiation 8, 495-505.
[13] Fritzsching, B., Schwer, B., Kartenbeck, J., Pedal, A.,
*Horejsi, V., Ott, M. 2002. Release and intercellular transfer of cell
surface CD81 via microparticles. J. Immunol. 169, 5531-5537.
[27] *Hollnagel, A., Grund, C., Franke, W.W., *Arnold, H.-H.
2002. The cell adhesion molecule M-cadherin is not essential for
muscle development and regeneration. Mol. Cell. Biol. 22, 47604770.
[28] Jave-Suarez, L.F., Winter, H., Langbein, L., Rogers, M.A.,
Schweizer, J. 2002. HoxC13 is involved in the regulation of human hair keratin gene expression. J. Biol. Chem.277, 3718-3726.
[29] *Kirfel, J., *Peters, B., Grund, C., *Reifenberg, K., *Magin,
T.M. 2002. Ectopic expression of desmin in the epidermis of
transgenic mice permits development of a normal epidermis. Differentiation 70, 56-68.
DKFZ 2003: Research Report 2001/2002
19
Research Program A
Cell Differentiation and Carcinogenesis
20
Division A0100
Cell Biology
[30] Kneissel, S. 2001. Ein neuartiges Karyoskelettprotein: Protein NO145, Hauptkomponente des nukleolären Cortex von Xenopus laevis Oocyten. Ph.D. Thesis. Faculty of Biology, University
of Heidelberg.
[45] *Reichelt, J., *Büssow, H., Grund, C., *Magin, T.M. 2001.
Formation of a normal epidermis supported by increased stability
of keratins 5 and 14 in keratin 10 null mice. Mol. Biol. Cell 12,
1557-1568.
[31] Kneissel, S., Franke, W.W., *Gall, J.G., Heid, H.,
Reidenbach, S., Schnölzer, M., Spring, H., Zentgraf, H.,
Schmidt-Zachmann, M.S. 2001. A novel karyoskeletal protein:
characterization of protein NO145, the major component of nucleolar cortical skeleton in Xenopus oocytes. Mol. Biol. Cell 12,
3904-3918.
[46] Reichenzeller, M. 2002. Strukturelle und dynamische
Analyse des Interchromosomalen Domänen-Kompartiments.
Ph.D. Thesis. Faculty of Biology, University of Heidelberg.
[32] *Kurzen, H., *Esposito, L., Langbein, L., *Hartschuh, W.
2001. Cytokeratins as markers of follicular differentiation. An immunohistochemical study of Trichoblastoma and basal cell carcinoma. Am. J. Dermatopathol. 23, 501-509.
[33] *Kurzen, H., *Manns, S., Dandekar, G., Schmidt, T.,
Prätzel, S., Kräling, B.M. 2002. Tightening of endothelial cell
contacts: a physiologic response to cocultures with smoothmusle-like 10T1/2 cells. J. Invest. Dermatol. 119, 143-153.
[34] Langbein, L., Rogers, M.A., Winter, H., Praetzel, S.,
Schweizer, J. 2001. The catalog of human hair keratins. II. Expression of the six type II members in the hair follicle and the
combined catalog of human type I and II keratins. J. Biol. Chem.
276, 35123-35132.
[35] Langbein, L., Grund, C., Kuhn, C., Praetzel, S.,
Kartenbeck, J., *Brandner, J.M., *Moll, I., Franke, W.W. 2002a.
Tight junctions and compositionally related junctional structures in
mammalian stratified epithelia and cell cultures derived therefrom. Eur. J. Cell Biol. 81, 419-435.
[36] Langbein, L., Rogers, M.A., Praetzel, S., *Aoki, N., Winter,
H., Schweizer, J. 2002b. A novel epithelial keratin, hK6irs1, is expressed differentially in all layers of the inner root sheath, including specialized Huxley cells (Flügelzellen) of the human hair follicle. J. Invest. Dermatol. 118, 789-799.
[37] Mertens, C., Hofmann, I., *Wang, Z., *Teichmann, M.,
*Sepehri Chong, S., Schnölzer, M., Franke, W.W. 2001. Nuclear
particles containing RNA polymerase III complexes associated
with the junctional plaque protein plakophilin 2. Proc. Natl. Acad.
Sci. USA 98, 7795-7800.
[38] *Möhrlen, F., *Baus, S., *Gruber, A., Rackwitz, H.-R.,
Schnölzer, M., *Vogt, G., *Zwilling, R. 2001. Activation of proastacin. Immunological and model peptide studies on the processing of immature astacin, a zinc-endopeptidase from the crayfish Astacus astacus. Eur. J. Biochem. 268, 2540-2546.
[39] *Olins, A.L., Herrmann, H., Lichter, P., *Kratzmeier, M.,
*Doenecke, D., *Olins, D.E. 2001. Nuclear envelope and chromatin compositional differences comparing undifferentiated and
retinoic acid- and phorbol ester-treated HL-60 cells. Exp. Cell
Res. 268, 115-127.
[40] Peitsch, W.K., Hofmann, I., Prätzel, S., Grund, C., Kuhn,
C., *Moll, I., Langbein, L., Franke, W.W. 2001.Drebrin particles:
components in the ensemble of proteins regulating actin dynamics of lamellipodia and filopodia. Eur. J. Cell Biol. 80, 567-579.
[41] Peitsch, W. 2002. Drebrin und Drebrosomen - Weit
verbreitete Zellkomponenten des Aktin-Cytoskelett-Systems. M.D.
Thesis, Faculty of Medicine, University of Hamburg.
[42] *Pelkmans, L., Kartenbeck, J., *Helenius, A. 2001.Caveolar
endocytosis of simian virus 40 reveals a new two-step vesiculartransport pathway to the ER. Nature Cell Biol. 3, 473-483.
[43] Pfrepper, K.-I., Reed, J., Rackwitz, H.-R., Schnölzer, M.,
Flügel, R.M. 2001. Characterization of peptide substrates and viral enzyme that affect the cleavage site specificity of the human
spumaretrovirus proteinase. Virus Genes 22, 61-72.
[44] Popanda, O., Flohr, C., Dai, J.-C., Hunzi(c)ker, A.,
Thielmann, H.W. 2001. A mutation in subunit B of the DNA polymerase α-primase complex from Novikoff hepatoma cells concomitant with a conformational change and abnormal catalytic
properties of the DNA polymerase α-primase complex. Mol. Carcinogenesis 31, 171-183.
[47] Rogers, M.A., Langbein, L., Winter, H., Ehmann, C.,
Praetzel, S., Korn, B., Schweizer, J. 2001. Characterization of a
cluster of human high/ultrahigh sulfur keratin-associated protein
genes embedded in the type I keratin gene domain on chromosome 17q12-21. J. Biol. Chem. 276, 19440-19451.
[48] #Rogers, M.A., #Langbein, L., Winter, H., Ehmann, C.,
Praetzel, S., Schweizer, J. 2002. Characterization of a first domain of human high glycine-tyrosine and high sulfur keratin-associated protein (KAP) genes on chromosome 21q22.1. J. Biol.
Chem. 277, 48993-49002. (#equal contribution)
[49] Rothbarth, K., Hunziker, A., Stammer, H., Werner, D. 2001.
Promoter of the gene encoding the 16 kDa DNA-binding and
apoptosis-inducing C1D protein. Biochim. Biophys. Acta 1518,
271-275.
[50] *Schaffeld, M., Herrmann, H., *Schultess, J., *Markl, J.
2001. Vimentin and desmin of a cartilaginous fish, the shark
Scyliorhinus stellaris: sequnece, expression patterns and in vitro
assembly. Eur. J. Cell Biol. 80, 692-702.
[51] Schumacher, J. 2001. Herstellung und Erprobung von
Lamin-Fragmenten in vitro und in vivo. Diploma Thesis. Faculty of
Biology, University of Heidelberg.
[52] *Shimomura, Y., *Aoki, N., Rogers, M.A., Langbein, L.,
Schweizer, J., *Ito, M. 2002a. hKAP1.6 and hKAP1.7, two novel
human high sulfur keratin-associated proteins are expressed in
the hair follicle cortex. J. Invest. Dermatol. 118, 226-231.
[53] *Shimomura, Y., *Aoki, N., Schweizer, J., Langbein, L.,
Rogers, M.A., Winter, H., *Ito, M. 2002b. Polymorphisms in the
human high sulfur hair keratin-associated protein 1, KAP1, gene
family. J. Biol. Chem. 277, 45493-45501.
[54] *Strelkov, S.V., Herrmann, H., *Geisler, N., *Lustig, A.,
*Ivaninskii, S., Zimbelmann, R., *Burkhard, P., *Aebi, U. 2001.
Divide-and-conquer crystallographic approach towards an atomic
structure of intermediate filaments. J. Mol. Biol. 306, 773-781.
[55] *Strelkov, S.V., Herrmann, H., *Geisler, N., Wedig, T.,
Zimbelmann, R., *Aebi, U., *Burkhard, P. 2002. Conserved segments 1A and 2B of the intermediate filament dimer: their atomic
structures and role in filament assembly. EMBO J. 21, 1255-1266.
[56] *Stumptner, C., *Fuchsbichler, A., Heid, H., *Zatloukal, K.,
*Denk, H. 2002. Mallory body - a disease-associated type of
sequestosome. Hepatology 35, 1053-1062.
[57] *Thai, T.P., *Rodemer, C., *Jauch, A., Hunziker, A., *Moser,
A., *Gorgas, K., *Just, W.W. 2001. Impaired membrane traffic in
defective ether lipid biosynthesis. Hum. Mol. Genet. 10, 127-136.
[58] *Volkmann, M., *Martin, L., *Bäurle, A., Heid, H.,
*Strassburg, C.P., *Trautwein, C., *Fiehn, W., *Manns, M.P. 2001.
Soluble liver antigen: Isolation of a 35-kd recombinant protein
(SLA-p35) specifically recognizing sera from patients with autoimmune hepatitis. Hepatology 33, 591-596.
[59] *Walther, T.C., *Pickersgill, H.S., Cordes, V.C., *Goldberg,
M.W., *Allen, T.D., *Mattaj, I.W., *Fornerod, M. 2002. The cytoplasmic filaments of the nuclear pore complex are dispensable for
selective nuclear protein import. J. Cell Biol. 158, 63-77.
[60] Winter, H., Langbein, L., *Krawczak, M., *Cooper, D.N.,
Jave-Suarez, L.F., Rogers, M.A., Praetzel, S., *Heidt, P.J.,
Schweizer, J. 2001. Human type I hair keratin pseudogene
ΨhHaA has functional orthologs in the chimpanzee and gorilla:
evidence for recent inactivation of the human gene after the PanHome divergence. Hum. Genet. 108, 37-42.
DKFZ 2003: Research Report 2001/2002
Research Program A
Cell Differentiation and Carcinogenesis
Division A0100
Cell Biology
[61] *Wolfrum, C., Borrmann, C.M., *Börchers, T., *Spener, F.
2001. Fatty acids and hypolipidemic drugs regulate peroxisome
prolifertor-activated receptors α- and γ-mediated gene expression
via liver fatty acid binding protein: a signaling path to the nucleus.
Proc. Natl. Acad. Sci. USA 98, 2323-2328.
Flow Cytometry Resource Group (A0102 /
A016)
[62] *Zatloukal, K., *Stumptner, C., *Fuchsbichler, A., Heid, H.,
Schnölzer, M., *Kenner, L., *Kleinert, R., *Prinz, M., *Aguzzi, A.,
*Denk, H. 2002. p62 is a common component of cytoplasmic inclusions in protein aggregation diseases. Am. J. Pathol. 160, 255263.
Tel: +49 - 6221 - 42 3208; FAX: +49 - 6221 - 42 2652
E-mail: [email protected]
Group leader: Michael Stöhr
Assistant: Monika Frank-Stöhr
Flow and image cytometry are two major approaches to
analytical cytology, which has been defined as the measurement and characterization of cells and cellular constituents for biological and medical purposes.
Flow cytometric analyses have been provided to various
research groups in the DKFZ in order to standardize the
setup and follow-up of their experiments.
Major activities of the group have been devoted to:
• service measurements, instruction, education, and advice on a routine basis;
• collaborations with scientific merit;
• instrumentation development with special emphasis on
personal computer data processing and analysis;
• organization of the annual Heidelberg Cytometry Symposium.
In detail, analytical duties referred to:
• analysis of cell cycle phases and degree of culture synchrony as well as apoptosis;
• analysis of DNA index and degree of polyploidy;
• analysis and electronic sorting of cells
• live-dead cell discrimination combined with estimation
of cell cycle phases;
• laser beam irradiation of cells incubated with photosensitive agents;
• quantitation of immunofluorescence (cell phenotyping).
We progressed in developing and expanding an x86 processor environment based on MS-DOS, the C programming language and some FORTRAN routines.
So, the flow cytometric data processing system is extremely versatile, user friendly, and user program-mable.
The current achievement comprises a complete 4 - parameter system for acquisition, real time display, storage,
retrieval, analysis, and documentation of cytometric data.
Additionally, a foreign file handler has been developed
and established to read cytometric data files from instruments of various manufacturers and convert the data to be
read from text and graphic processing programs under
Windows 3.xx and higher.
Thus, flow cytometric data can be integrated into text processing routines as doc, bmp, pcx, tif or other file formats.
These facilities provide access to cytometry instrumentation, are operative on a service basis, supply instructions
for independent and easy use of cytometry equipment and
to all conflicts and questions of cytometry in general, and
are intended to develop new techniques and expand the
area of cytometry applications in biomedical research at
DKFZ.
DKFZ 2003: Research Report 2001/2002
21