Role of GAS2L3 in cardiomyocyte proliferation and cytokinesis in vitro and in vivo

• Applicant: Professor Dr. Stefan Gaubatz
• Subject Area: Cell Biology ; Developmental Biology
• Term: from 2015 to 2022
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 288877080

The failure to undergo cytokinesis is one of the limiting steps that prevents heart regeneration after injury. A better understanding of the mechanisms that control cardiac cell cycle progression and cytokinesis is therefore necessary to develop new therapies for heart diseases. We have recently identified GAS2L3, a target gene of the DREAM/ B-MYB multi-protein complex, as a novel protein involved in cytokinesis. GAS2L3 acts as a mitotic cytolinker protein that can cross-link actin filaments with the microtubule network. To investigate the in vivo function of GAS2L3 we have created a knockout mouse model and found that Gas2l3 k.o. mice die postnatally from dilated cardiomyopathy. Further studies suggest that GAS2L3 functions in cardiomyocyte cytokinesis during the critical postnatal period when cardiomyocytes binucleate and exit from the cell cycle. The main objective of this proposal is to expand on these results to gain insights into the role of GAS2L3 in cardiomyocyte cell cycle regulation. Our conditional and non-conditional Gas2l3 knockout mouse lines will be used in this study. In the first part of the proposal, we will investigate the role of Gas2l3 in cardiomyocyte proliferation and cytokinesis by immunohistochemistry. We will also perform live-cell imaging of conditional Gas2l3 knockout cardiomyocytes expressing fluorescent fusion proteins to monitor cell division and cytokinesis. In the second part of the proposal we will test the hypothesis that GAS2L3 acts to coordinate the breakdown of the contractile apparatus with cytokinesis. To address this possibility we will investigate the biochemical interaction of GAS2L3 with Z-disc proteins and cytokinesis proteins and analyze the functional significance of these interactions. We will also analyze the sarcomere structure and myofibril disassembly of Gas2l3 k.o. cardiomyocytes. In the final part of the proposal we will test the hypothesis that DREAM is responsible for the coordinated expression of Gas2l3 and other cytokinesis genes during cardiomyocyte development. To directly test the role of DREAM in cardiomyocyte development we will generate mice with cardiac-specific deletion of DREAM subunits. To identify direct targets of DREAM in cardiomyocytes we will perform RT-qPCR, RNA-Seq and ChIP-Seq experiments. We will also investigate a possible crosstalk between Hippo signaling and the DREAM complex in gene expression in cardiomyocytes.Overall our studies will provide new insights into the mechanisms of binucleation and cell cycle withdrawal of cardiomyocytes.

• DFG Programme: Research Grants