Graduation Year

2021

Document Type

Dissertation

Degree

Ph.D.

Degree Name

Doctor of Philosophy (Ph.D.)

Degree Granting Department

Biology (Cell Biology, Microbiology, Molecular Biology)

Major Professor

Sandy D. Westerheide, Ph.D.

Committee Member

Meera Nanjundan, Ph.D.

Committee Member

Younghoon Kee, Ph.D.

Committee Member

Margaret Park, Ph.D.

Committee Member

Eric Guisbert, Ph.D.

Keywords

RNA, heat shock, stress response, UAF-1

Abstract

The Cell Division Cycle and Apoptosis Regulator (CCAR) family members are an enigmatic family of proteins regulating metabolism, cancer, apoptosis, DNA damage, and stress. Mammals have CCAR family members, CCAR1 and CCAR2/DBC1, which evolved from the founding family member CCAR-1/LST-3 expressed in Caenorhabditis elegans. Several studies have shown the importance of understanding these proteins' function in standard and altered physiological processes. Our studies aim to understand the genome-wide alternative splicing and germline regulation of Caenorhabditis elegans CCAR-1 in normal and heat shock conditions. Recently, mammalian CCAR family member CCAR2/DBC1 regulates the alternative splicing by forming a complex with ZNF326. This complex bind both RNA polymerase II and the ribonucleoprotein particle, integrating alternative splicing with RNA polymerase II elongation in A+T regions of the DNA. Additionally, C. elegans CCAR-1 affects the perlecan gene unc-52 alternative splicing by allowing the excision of exon 17 during development. However, the CCAR-1 genome-wide alternative splicing regulation in vivo and during stress remains unknown. Also, the mechanism of action of this regulation is yet to be understood. Using RNA sequencing, we identify new alternative splicing targets with the deletion of CCAR-1. Furthermore, through mass spectrometry, we show the interaction of CCAR-1 with splicing factors of the spliceosome UAF-1 and UAF-2, unraveling a potential mechanism of action of CCAR-1. Also, our whole transcriptome RNA sequencing identifies CCAR-1’s role in the gene expression of Caenorhabditis elegans. Our analysis shows that CCAR-1 regulates germline transcription, reproduction, lifespan, and DNA-damage-induced apoptosis, a role that is previously unknown to the CCAR family. Overall, this work increases our knowledge of the CCAR family’s role in alternative splicing and identifies a new function in the C. elegans germline.

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