Thesis (MSc)--University of Stellenbosch, 2003. / ENGLISH ABSTRACT: Yeasts, like most organisms, have to survive in highly variable and hostile
environments. Survival therefore requires adaptation to the changing external
conditions. On the molecular level, specific adaptation to specific environmental
conditions requires the yeast to be able: (i) to sense all relevant environmental
parameters; (ii) to relay the perceived signals to the interior of the cell via signal
transduction networks; and (iii) to implement a specific molecular response by
modifying enzyme activities and by regulating transcription of the appropriate genes.
The availability of nutrients is one of the major trophic factors for all unicellular
organisms, including yeast. Saccharomyces cerevisiae senses the nutritional
composition of the media and implements a specific developmental choice in response
to the level of essential nutrients. In conditions in which ample nutrients are available,
S. cerevisiae will divide mitotically and populate the growth environment. If the
nutrients are exhausted, diploid S. cerevisiae cells can undergo meiosis, which
produces four ascospores encased in an ascus. These ascospores are robust and
provide the yeast with a means to survive adverse environmental conditions. The
ascospores can lie dormant for extended periods of time until the onset of favourable
growth conditions, upon which the spores will germinate, mate and give rise to a new
yeast population. However, S. cerevisiae has a third developmental option, referred to
as pseudohyphal and invasive growth. In growth conditions in which nutrients are
limited, but not exhausted, the yeast can undergo a morphological switch, altering its
budding pattern and forming chains of elongated cells that can penetrate the growth
substrate to forage for nutrients.
The focus of this study was on elements of the signal transduction networks
regulating invasive growth in S. cerevisiae. Some components of the signal
transduction pathways are well characterised, while several transcription factors that
are regulated via these pathways remain poorly studied. In this study, the RMEt gene
was identified for its ability to enhance starch degradation and invasive growth when
present on a multiple copy plasmid. Rme1 p had previously been identified as a
repressor of meiosis and, for this reason, the literature review focuses on the
regulation of the meiotic process. In particular, the review focuses on the factors
governing entry into meiosis in response to nutrient starvation and ploidy. Also, the
transcriptional regulation of the master initiator of meiosis, IMEt, and the action of
Ime1 p are included in the review.
The experimental part of the study entailed a genetic analysis of the role of Rme1 p
in invasive growth and starch metabolism. Epistasis analysis was conducted of
Rme1 p and elements of the MAP Kinase module, as well as of the transcription
factors, Mss11p, Msn1p/Mss10p, Tec1p, Phd1p and F108p. Rme1p is known to bind
to the promoter of CLN2, a G1-cyclin, and enhances its expression. Therefore, the cell cyclins CLN1 and CLN2 were included in the study. The study revealed that Rme1 p
functions independently or downstream of the MAP Kinase cascade and does not
require Cln1 p or Cln2p to induce invasive growth. FL011/MUC1 encodes a cell wall
protein that is required for invasive growth. Like the above-mentioned factors, Rme1 p
requires FL011 to induce invasive growth. We identified an Rme1 p binding site in the
promoter of FL011. Overexpression of Rme1p was able to induce FL01t expression,
despite deletions of mss11, msn1, ttos, tee1 and phd1. In the inverse experiment,
these factors were able to induce FL011 expression in an rme1 deleted strain. This
would indicate that Rme1 p does not function in a hierarchical signalling system with
these factors, but could function in a more general role to modify transcription. / AFRIKAANSE OPSOMMING: Die natuur is hoogs veranderlik en alle organismes, insluitende gis, moet by die
omgewing kan aanpas om te kan oorleef. Baie eksterne faktore beïnvloed die
ontwikkeling van die gissel. Vir die gis om by spesifieke omgewingstoestande aan te
pas, moet die gis op 'n molekulêre vlak: (i) al die omgewingsparameters waarneem; (ii)
die waargenome omgewingsparameters as seine na die selkern deur middel van
seintransduksieweë gelei; en (iii) transkripsie van gene aktiveer of onderdruk en
ensiemaktiwiteit reguleer om sodoende die gepaste molekulêre respons te
implementeer.
Die beskikbaarheid van voedingstowwe in die omgewing is een van die
belangrikste omgewingseine wat eensellige organismes moet kan waarneem.
Saccharomyces cerevisiae kan spesifieke ontwikkelingsopsies, na gelang van die
voedingstowwe wat beskikbaar is, uitoefen. In groeiomstandighede waar daar 'n
oorvloed van voedingstowwe is, verdeel S. cerevisiae d.m.v. mitose en vesprei dit
deur die omgewing. Sodra die voedingstowwe uitgeput is, word mitose onderdruk.
Diploïede S. cerevisiae inisieer meiose, wat aanleiding tot die vorming van vier spore
gee. Hierdie spore bevat slegs die helfte van die ouer se chromosome en kan
gevolglik met 'n ander spoor paar om weer 'n diploïede gissel te vorm. Die spore is
bestand teen strawwe omgewingstoestande en kan vir lang tye oorleef. Wanneer die
spoor aan gunstige groeitoestande blootgestel word, ontkiem dit om aan 'n nuwe
giskolonie oorsprong te gee. S. cerevisiae het egter 'n derde ontwikkelingsopsie,
naamlik pseudohife-differensiëring. Wanneer die beskikbaarheid van voedingstowwe
in die omgewing afneem, maar nog nie uitgeput is nie, ondergaan die gis 'n
morfologiese verandering. Hierdie verandering word gekenmerk deur selverlenging,
nl. botselle wat slegs aan die een punt van die gissel vorm en dogterselle wat aan die
moerderselle geheg bly. Dit lei tot die vorming van kettings van selle wat van die
giskolonie af weggroei. Voorts kan die selkettings ook die groeisubstraat binnedring.
Dit staan as penetrasie-groei bekend en laat die gis toe om na nuwe voedingsbronne
te soek.
Hierdie studie het op die elemente van seintransduksieweë, wat by
penetrasiegroei betrokke is, gefokus. Sekere komponente van die seintransduksieweë
is reeds goed gekarakteriseer, terwyl ander komponente nog grootliks onbekend is. In
hierdie studie, word 'n rol vir RME1 in die verbetering van styselafbraak en
penetrasiegroei geïdentifiseer. Aangesien Rme1 p voorheen as 'n onderdrukker van
meiose geïdentifiseer is, is 'n litetaruurstudie oor die inisiasie van meiose saamgestel.
Die faktore wat meiose induseer, naamlik 'n gebrek aan voedingstowwe en die sel se
ploïedie, word bespreek. Die regulering van die meester inisieerder van meiosie,
IME1, asook die proteïene waarmee Ime1p reageer, is ook in die studie ingesluit. Die eksperimentele deel van die studie behels die genetiese analise van Rme1 p
tydens penetrasiegroei en styselhidroliese. 'n Epistase-analise tussen Rme1 p en
elemente van die MAP-Kinasemodule, asook van die transkripsie faktore Mss11 p,
Msn1p/Mss10p, Tec1p, Phd1p en F108p, is onderneem. Rme1p is bekend om aan die
promotor van CLN2 te bind en transkripsie te induseer. Daarom is die selsikliene
CLN1 en CLN2 in die studie ingesluit. Die studie dui daarop dat Rme1 ponafhanklik
van die MAP-Kinasemodule funksioneer en nie Cln1 p en Cln2p benodig om
penetrasiegroei te induseer nie. FL011/MUC1 kodeer vir 'n selwandproteïen wat
noodsaaklik vir pentrasiegroei is. Soos in die geval van die bogenoemde faktore,
benodig Rme1 p FL011 om penetrasiegroei te kan induseer. Ten spyte van mss11-,
msn1-, ttos-, tec1- en phd1- delesies, kan ooruitdrukking van Rme1p die transkripsie
van FL011 induseer. In die omgekeerde eksperiment kon die bogenoemde faktore
FL011-transkripsie ten spyte van 'n rme1 delesie induseer. Die resultate dui daarop
dat Rme1 p nie in 'n hiërargiese pad funksioneer nie, maar dat dit waarskynlik 'n meer
algemene rol deur transkripsiemodifisering vervul.
| Identifer | oai:union.ndltd.org:netd.ac.za/oai:union.ndltd.org:sun/oai:scholar.sun.ac.za:10019.1/53322 |
| Date | 03 1900 |
| Creators | Hansson, Guy Robert, 1974- |
| Contributors | Bauer, F. F., Pretorius, I. S., Stellenbosch University. Faculty of AgriSciences. Dept. of Viticulture and Oenology. Institute for Wine Biotechnology. |
| Publisher | Stellenbosch : Stellenbosch University |
| Source Sets | South African National ETD Portal |
| Language | en_ZA |
| Detected Language | Unknown |
| Type | Thesis |
| Format | 72 p. : ill. |
| Rights | Stellenbosch University |
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