Zur Seitenansicht
 

Titelaufnahme

Titel
Elucidating the LIM-Code in pancreatic endocrine cells / Author: Fabian Martin
VerfasserMartin, Fabian
Betreuer / BetreuerinnenMeyer, Dirk
ErschienenInnsbruck, 2018
UmfangII, 62 Blätter : Illustrationen, Diagramme
HochschulschriftUniversity Innsbruck, Masterarbeit, 2018
Datum der AbgabeJuni 2018
SpracheEnglisch
DokumenttypMasterarbeit
Schlagwörter (DE)LIM-Code / Pancreas / Islet1
URNurn:nbn:at:at-ubi:1-24042 Persistent Identifier (URN)
Zugriffsbeschränkung
 Das Werk ist frei verfügbar
Dateien
Elucidating the LIM-Code in pancreatic endocrine cells [1.88 mb]
Links
Nachweis
Klassifikation
Zusammenfassung (Englisch)

The “LIM-Code” describes a model for generating distinct neuronal cell types by combinatorial function of distinct sets of LIM-domain transcription factors in an initially similar progenitor population. Support for this model comes from studies in C. elegans, Drosophila and various vertebrates, which demonstrated cell-type-specific requirements for LIM-proteins and their interaction during differentiation, maturation and survival of various neurons. In this Master project, I focused on pancreatic LIM-protein function to determine a potential pancreatic LIM-code. Previous work in our group established that in zebrafish (D. rerio) LIM-homeobox factor Islet1 (isl1) has cell-type specific functions in the developing pancreatic islet and that these distinct functions correlate with different expression profiles of several LIM-proteins encoding genes. To allow more detailed analyses of the cell-type specific isl1-requirements, a genome editing approach to replace the first exons of isl1 in zebrafish was initiated. A corresponding targeting vector for homology directed repair mutagenesis was generated that in exon 1 contained additional sequences for a fluorescent protein and a viral epitope tag and in which exon 2 was flanked by LoxP sites. The idea of this construct was to bypass postembryonic lethality of isl1 by using conditional and cell-type specific gene inactivation and to use these lines for in vivo imaging of isl1-expression and in ChIP approached using the epitope.

Further, detailed molecular analyses of rat cell lines resembling characteristics of pancreatic - and -cells showed that co-expression of isl1 with a cell-type-specific LIM-factors was not restricted to zebrafish. Using the rat cell-lines as a model to study the basis of a pancreatic LIM-code confirmed direct interaction between these proteins and I established protocols to purify and analyse additional co-factors by mass spectrometry.

In addition, CRISPR/Cas9 tools for genome editing were established to mutate Isl1 in the pancreatic rat cell lines. While these tools induced mutations, corresponding Isl1-deficient cell lines could not be recovered. The data suggest that isl1 is essential for survival or growth of these cells.

Statistik
Das PDF-Dokument wurde 10 mal heruntergeladen.