טכניון מכון טכנולוגי לישראל
הטכניון מכון טכנולוגי לישראל - בית הספר ללימודי מוסמכים  
M.Sc Thesis
M.Sc StudentKheir Sally
SubjectUnmasking RegulatoryPpaths Involved in the Setting of
Animal Cell Fates
DepartmentDepartment of Biology
Supervisor Professor Itai Yanai
Full Thesis textFull thesis text - English Version


Abstract

The astounding cell diversity of an animal arises from the ability of each cell in the developing embryo to commit to a particular fate. A cell’s ‘competence’ relates to its ability to respond to specific signals and differentiate into a particular fate. Importantly, this state of competence is limited to stages in development, beyond which the cell’s fate are not alterable. Thus, 'competence' in this sense means the ability of a cell to become reprogrammed as a response to a set of specific signals. However, the molecular mechanisms underlying the opening and the closing of a competence window are not well understood. In this work, I study the precise gene expression states as the fate of an individual C.elegans blastomere is reprogrammed. The ‘E blastomere specifies the endoderm. However, previous work has shown that induction of the MyoD ortholog (hlh-1) can reprogram this to mesoderm during a particular competence window. We used CEL-Seq, a multiplexed method for single -cell RNA-Seq, to examine the molecular changes that occur in cultured E blastomeres with exogenously introduced MyoD at different developmental stages. Surprisingly, we observed that after the gut fate is specified in the E cell, induction of MyoD at the two-cell stage of E, resulted in a muscle fate conversion in only 50% of the E cells, with the remaining continuing to develop to gut. This result suggests an opening in the competence window at this stage. I next investigated how hlh-1 interacts with the endoderm fate to decipher the basis of cell fate acquisition and competence by establishing a developmental time-course of induced E blastomeres in its competent state. Our results show that the transition to a new fate induced by an extrinsic cue is a gradual process that first requires the elimination of the original specification program before the initiation of the new specification program. We hypothesize that the competence window serves as a transition between specification of a fate and its full differentiation. We find that cell fates cannot change during the specification step but are amenable to change during the transition or differentiation steps and propose that chromatin regulators might play a central role in this process . Termination of the competence window occurs once the differentiation cascade reaches a threshold and stabilizes a specific fate. Hence, we envision the 'competence window' as the central element of cell fate decisions. Unlocking the dynamics of developmental plasticity can also introduce evolutionary insights. We found that cells with different fates respond differently to MyoD signaling. The ectoderm specifying AB blastomere, still retained vestiges of its AB-identity following MyoD induction and the appearance of mesoderm marker gene expression. This result may be explained by the evolutionary history of the germ layers: if the mesoderm evolved from the endoderm as has been proposed, then the endoderm may be more amenable to change than the ectoderm because of its closer evolutionary history. Our results provide a detailed time-line of cell fate conversion and highlight the constraints which the ancestry of the germ layers imposes on the development of the organism . _____________________________________________________________________________