Regulation of binding of HP1 associated complexes to chromatin and their role in transcription regulation in C. elegans vulva development

by Yogesh Ostwal
Doctoral thesis
Date of Examination:2105-10-21
Date of issue:2016-10-27
Advisor:Prof. Dr. Wolfgang Fischle
Referee:Prof. Dr. Herbert Jäckle
Referee:Prof. Dr. Michael Kessel
Referee:Dr. Dieter Klopfenstein
Persistent Address: http://dx.doi.org/10.53846/goediss-5901

 

 

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Abstract

English

HP1 proteins are known to bind to H3K9me3 and thereby localize to heterochromatin. However, direct functions of HP1 in heterochromatin formation and maintenance are not clear. This is partially due to essentiality of HP1 proteins in mammals and flies. We therefore, explored the functions of HP-1 homologues in C. elegans. Our findings suggest that the C. elegans HP1 homologue; HPL-2 does not directly bind H3K9me3, unlike what has been proposed for HP1 family proteins. Interestingly, HPL-2 depends on LIN-61 for association with H3K9me3. Our analysis of LIN-61-H3K9me3 interaction defective mutant suggests that HPL-2-LIN-61-H3K9me3 interaction is required for vulva development in C. elegans. We further demonstrate that HPL-2, LIN-61 and LIN-13 biochemically interact with each other to form multiple complexes. We show that HPL-2 interacts with both LIN-61 and LIN-13 via its CSD. Moreover, we show that LIN-61 and LIN-13 do not directly interact with each other in the absence of HPL-2. Consequently, full length HPL-2 and possibly its multimerization is required for LIN-61-HPL-2-LIN-13 complex formation. Our results suggest existence of at least three putative complexes among HPL-2, LIN-13 and LIN-61. These complexes bind distinct, non-overlapping subsets of target genes and regulate their transcription including genes associated with vulva development. In agreement, hpl-2, lin-61 and lin-13 genetically interact with each other during vulva development. Analysis of the binding sites of HPL-2-LIN-61 complex and HPL-2-LIN-13 complex reveals two distinct modes of chromatin association of HPL-2. Sharp distribution of HPL-2-LIN-13 complex points to transcription factor mediated recruitment. Broad distribution of HPL-2-LIN-61 complex points to ncRNA mediated recruitment. Our results on involvement of a RNA component in the stability of HPL-2-LIN-61-LIN13 complexes reiterate our findings. Overall, our studies highlight the role of HP1 associated complexes in transcription regulation in vulva development in C. elegans.
Keywords: HP1; C. elegans; Heterochromatin; HPL-2; Vulva development
 
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