ZMBH J.Bericht 2000 - Zentrum für Molekulare Biologie der ...

ferential display methods and gene arrays have been
initiated and putative candidates are currently analyzed.
Pseudohyphal and hyphal growth plays an important
role during pathogenesis of the yeast Candida albicans.
In the course of the C. albicans sequencing project,
a DNA fragment of the putative C. albicans ASH1
gene has been identified. Using this fragment, we have
cloned the full length C. albicans ASH1 gene and are
currently characterizing its function.
Novel localized mRNAs in Saccharomyces
cerevisiae
F. Böhl and D. Ferring
In a number of cell types, several mRNAs are sorted
differentially in a single cell. In order to identify novel
localized RNAs in yeast, we devised two different
screens based on in-situ hybridization. The first one
was based on the rationale that asymmetrically sorted
mRNAs should be expressed in the same cell cycle
window as ASH1, the M-/G1-phase boundary. Therefore,
we fused the 3‘ parts of 40 mRNAs that are expressed
at this boundary to a reporter mRNA (GFP,
green fluorescent protein) and expressed these hybrids
from a regulatable promoter. In the case of ASH1 such
an RNA hybrid is clearly localized to the daughter cell.
However, none of the other mRNA hybrids showed a
localized hybridization signal, indicating that ASH1 is
the only M-/G1-phase mRNA that is asymmetrically
distributed.
In a second approach, we tempted to screen the complete
yeast genome for localized mRNAs. A plasmidbased
DNA library was generated that allowed random
fusions of genomic DNA fragments to the GFP coding
sequence. The resulting hybrid genes can be expressed
from a regulatable GAL1 promoter. Localized RNAs
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can be detected by in-situ hybridization against the
GFP part of the resulting hybrid mRNAs. The constructed
library covers approximately 95% of the complete
yeast genome. After transformation into yeast,
pools of 25 clones are currently analyzed for the presence
of novel localized mRNAs. Such a screen should
be useful to assess the complete number of localized
mRNAs in a polarized cell.
External Funding
During the period reported our research was supported
by grants from the Deutsche Forschungsgemeinschaft
DFG (Graduiertenkolleg “Molekulare Zellbiologie”,
Projekt-’Sachbeihilfen’).
PUBLICATIONS
Münchow, S., Sauter, C., and Jansen, R.-P. (1999). Association
of the class V myosin Myo4p with a localised
messenger RNA in budding yeast depends on She proteins.
J. Cell Sci. 112, 1511-1518.
Jansen, R.-P. (1999). RNA-cytoskeletal associations.
FASEB J. 13, 455-466.
Hurt, E., Segref, A., Sträßer, K., Bailer, S., Schlaich,
N., Presutti, C., Tollervey, D., and Jansen, R.-P. (2000).
Mex67p mediates nuclear export of a variety of Pol II
transcripts. J.Biol.Chem. 275, 8361-8368.
THESES
Diploma
Münchow, S. (1998). Isolierung und Charakterisierung
des Myo4-Protein/ASH1 mRNA Komplexes aus Sac-
charomyces cerevisiae.
Sauter, C. (1998). Detektion lokalisierter mRNAs in
Saccharomyces cerevisiae mittels Fluoreszenz-in situ-
Hybridisierung.
Theurer, J. (1998). Versuch der Identifikation neuer
Bestandteile der ASH1 mRNA Lokalisationsmaschi–
nerie von Saccharomyces cerevisiae durch Überexpression
einer cDNA-Bibliothek.
Djandji, D. (1999). Die Assoziation des She3 Proteins
mit ASH1 mRNA in Saccharomyces cerevisiae.
STRUCTURE OF THE GROUP
e-mail: r.jansen@mail.zmbh.uni-heidelberg.de
Group leader Jansen, Ralf-Peter, Dr.
Ph.D. students Böhl, Florian, Dipl. Biol. *
Jaedicke, Andreas, Dipl. Biol. *
Kruse, Claudia, Dipl. Biol. *
Münchow, Sonja, Dipl. Biol.
Diploma students Djandji, Dominic *
Sauter, Claus *
Theurer, Jörg *
Kahlina, Kornelia *
Techn. assistants Ferring, Dunja *
Frank, Andrea *
* part of the time reported
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