Identification of hcf-1 in a genetic screen for dsh-2 suppressors in c. Elegans

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(Thesis) Ph.D.
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Asymmetric cell division is an essential process to generate cell diversity during development. In C. elegans, many asymmetric cell divisions are regulated by Wnt signalling. We have identified a Wnt/CWN-1, a Frizzled/MOM-5 and a Dishevelled/DSH-2 that function to control asymmetric neuroblast division. Loss of both maternal and zygotic dsh-2 function results in asymmetric neuroblast division defects and embryonic/early larval lethality, while loss of zygotic dsh-2 function disrupts asymmetric cell division of the somatic gonadal precursor cells (SGPs), Z1 and Z4. Through a DSH-2 domain analysis, we found that the DEP domain of DSH-2 is required for viability and asymmetric neuroblast division, while neither the DIX or DEP domains were essential for SGP cell division. To identify genes that function with dsh-2 in asymmetric division, we undertook a genetic screen to isolate suppressors of dsh-2 lethality and isolated over 60 dominant suppressors. We focused our characterization on Sup305 and Sup245 which we demonstrated were also strong suppressors of both asymmetric neuroblast and SGP division defects. In SGP division, reciprocal asymmetric localization of SYS-1/β-catenin and POP-1/TCF in Z1/Z4 daughter cells regulate correct cell fate. This asymmetric localization is lost in dsh-2 mutants. Both suppressors partially restored the asymmetric localization of SYS-1/β-catenin and POP-1/TCF in dsh-2 mutants. Genetic mapping experiments placed Sup305 on the middle of chromosome IV and Sup245 on the right arm of chromosome I. Both suppressor strains were sent for whole genome sequencing and the resulting sequence analyzed to identify potential candidates. In combination with additional mapping experiments, we determined that Sup305 was a G to A in mutation in hcf-1 resulting in a Proline to Serine amino acid substitution. Loss of hcf-1 rescued dsh-2 phenotypes suggesting that Sup305 is a dominant negative mutation in hcf-1. hcf-1 is a transcriptional cofactor that bridges transcription factors to the chromatin modifying machinery. In C. elegans, hcf-1 has been previously shown to modulate cell cycle, stress and lifespan. Our results indicate that it is also a novel Wnt pathway interactor. Further work will determine the mechanism of hcf-1 suppression of dsh-2.
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Thesis advisor: Hawkins, Nancy
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