{"id":{"repo_id":"toronto-retro","oai_identifier":"oai:utoronto.scholaris.ca:1807/150378"},"canonical_url":"https://search.dev.ndltd.org/etd/toronto-retro/oai:utoronto.scholaris.ca:1807/150378","repository":{"repo_id":"toronto-retro","name":"University of Toronto","base_url":"https://utoronto.scholaris.ca/server/oai/request"},"display":{"title":"Uncovering the Roles of Human C2H2-Zinc Finger Protein ZNF121 in Gene Regulation","abstract":"Gene expression is a complex, dynamic, and tightly orchestrated process. Regulation of genes transcribed by RNA polymerase II is coordinated by an elegant arrangement of protein machinery and chromatin architecture, and the resulting pre-mRNA is post-transcriptionally processed and regulated at every step to buffer, reduce, or amplify its protein output. Chemical nucleotide modifications on DNA or RNA are one type of gene regulatory mechanism. N6-methyladenosine (m6A) is the most abundant and well-characterized mRNA modification, facilitating mRNA regulatory processes that include mRNA decay regulated by the m6A reader proteins YTH-domain-containing family protein 2 (YTHDF2). However, YTHDF2 has a relatively weak affinity for its mRNA targets. RNA-binding proteins (RBPs) bind transcripts to exert regulatory effects, and many DNA-binding C2H2-zinc finger proteins (ZFPs) have recently been shown to be RBPs with roles in mRNA biology. I identified C2H2-ZFP, ZNF121, as a YTHDF2 binding partner in affinity purification and mass-spectrometry (AP-MS) experiments and I hypothesized that, like other tested ZFPs, ZNF121 is an RNA-binding protein and acts as a cofactor to stabilize YTHDF2 on shared mRNA targets. In this thesis, I utilize multi-omic and biochemical approaches to understand the role of ZNF121 in gene expression. I observe that ZNF121 is a cyto-nuclear protein and confirm its interaction with YTHDF2 and RNA. Using iCLIPseq, I found that ZNF121 binds mRNA around sites of YTHDF2 enrichment, even when no nearby m6A modification exists. Specifically, over 80% of ZNF121-bound transcripts are also YTHDF2-bound transcripts. Upon depletion of ZNF121, the binding of YTHDF2 to shared transcripts is impaired, and the stability of these transcripts increases in an m6A-independent fashion. This affects many transcripts coding for cell-cycle related proteins like MDM2, with effects on the DNA damage response and cell viability. Lastly, I observe that ZNF121 regulates cassette exon retention in the nucleus. My results show that ZNF121, beyond its conventional role as a transcription factor, is a versatile protein that regulates both mRNA stability in conjunction with YTHDF2 and mRNA splicing.","abstract_html":"Gene expression is a complex, dynamic, and tightly orchestrated process. Regulation of genes transcribed by RNA polymerase II is coordinated by an elegant arrangement of protein machinery and chromatin architecture, and the resulting pre-mRNA is post-transcriptionally processed and regulated at every step to buffer, reduce, or amplify its protein output. Chemical nucleotide modifications on DNA or RNA are one type of gene regulatory mechanism. N6-methyladenosine (m6A) is the most abundant and well-characterized mRNA modification, facilitating mRNA regulatory processes that include mRNA decay regulated by the m6A reader proteins YTH-domain-containing family protein 2 (YTHDF2). However, YTHDF2 has a relatively weak affinity for its mRNA targets. RNA-binding proteins (RBPs) bind transcripts to exert regulatory effects, and many DNA-binding C2H2-zinc finger proteins (ZFPs) have recently been shown to be RBPs with roles in mRNA biology. I identified C2H2-ZFP, ZNF121, as a YTHDF2 binding partner in affinity purification and mass-spectrometry (AP-MS) experiments and I hypothesized that, like other tested ZFPs, ZNF121 is an RNA-binding protein and acts as a cofactor to stabilize YTHDF2 on shared mRNA targets. In this thesis, I utilize multi-omic and biochemical approaches to understand the role of ZNF121 in gene expression. I observe that ZNF121 is a cyto-nuclear protein and confirm its interaction with YTHDF2 and RNA. Using iCLIPseq, I found that ZNF121 binds mRNA around sites of YTHDF2 enrichment, even when no nearby m6A modification exists. Specifically, over 80% of ZNF121-bound transcripts are also YTHDF2-bound transcripts. Upon depletion of ZNF121, the binding of YTHDF2 to shared transcripts is impaired, and the stability of these transcripts increases in an m6A-independent fashion. This affects many transcripts coding for cell-cycle related proteins like MDM2, with effects on the DNA damage response and cell viability. Lastly, I observe that ZNF121 regulates cassette exon retention in the nucleus. My results show that ZNF121, beyond its conventional role as a transcription factor, is a versatile protein that regulates both mRNA stability in conjunction with YTHDF2 and mRNA splicing.","abstract_has_math":false,"creators":["Burke, Giovanni Livingston"],"institution":null,"degree_name":null,"degree_level":null,"degree_discipline":null,"degree_department":"Molecular Genetics","school":null,"contributors":[],"advisors":["Greenblatt, Jack F"],"committee_chairs":[],"committee_members":[],"year":2025,"date_issued":"2025-10","date_published":"2025-10","updated_at":"2026-07-27T21:28:20Z","subjects":["C2H2 Zinc Finger Protein","m6A","RNA biology","RNA modifications","YTHDF2","ZNF121"],"languages":[],"rights":["Attribution 4.0 International"],"rights_urls":["http://creativecommons.org/licenses/by/4.0/"],"identifier_entries":[]},"links":{"outbound_url":"https://hdl.handle.net/1807/150378","outbound_label":"Handle","outbound_source":"dc:identifier.uri"},"metadata_groups":[{"id":"people","label":"People","entries":[{"key":"dc:contributor.advisor","label":"Advisor","values":["Greenblatt, Jack F"]},{"key":"dc:contributor.department","label":"Department","values":["Molecular Genetics"]},{"key":"dc:creator","label":"Author","values":["Burke, Giovanni Livingston"]}]},{"id":"academic_context","label":"Academic Context","entries":[{"key":"dc:date","label":"Dc Date","values":["2025-10"]},{"key":"dc:date.accessioned","label":"Dc Date Accessioned","values":["2025-12-01T16:13:48Z"]},{"key":"dc:date.issued","label":"Date","values":["2025-10"]},{"key":"dc:type","label":"Dc Type","values":["Thesis"]}]},{"id":"subjects_keywords","label":"Subjects and Keywords","entries":[{"key":"dc:subject","label":"Dc Subject","values":["C2H2 Zinc Finger Protein","m6A","RNA biology","RNA modifications","YTHDF2","ZNF121"]}]},{"id":"language_rights","label":"Language and Rights","entries":[{"key":"dc:rights","label":"Dc Rights","values":["Attribution 4.0 International"]},{"key":"dc:rights.uri","label":"Rights URI","values":["http://creativecommons.org/licenses/by/4.0/"]}]},{"id":"identifiers","label":"Identifiers","entries":[{"key":"dc:identifier.uri","label":"Identifier URI","values":["https://hdl.handle.net/1807/150378"]}]},{"id":"additional","label":"Additional Metadata","entries":[{"key":"dc:description.abstract","label":"Abstract","values":["Gene expression is a complex, dynamic, and tightly orchestrated process. Regulation of genes transcribed by RNA polymerase II is coordinated by an elegant arrangement of protein machinery and chromatin architecture, and the resulting pre-mRNA is post-transcriptionally processed and regulated at every step to buffer, reduce, or amplify its protein output. Chemical nucleotide modifications on DNA or RNA are one type of gene regulatory mechanism. N6-methyladenosine (m6A) is the most abundant and well-characterized mRNA modification, facilitating mRNA regulatory processes that include mRNA decay regulated by the m6A reader proteins YTH-domain-containing family protein 2 (YTHDF2). However, YTHDF2 has a relatively weak affinity for its mRNA targets. RNA-binding proteins (RBPs) bind transcripts to exert regulatory effects, and many DNA-binding C2H2-zinc finger proteins (ZFPs) have recently been shown to be RBPs with roles in mRNA biology. I identified C2H2-ZFP, ZNF121, as a YTHDF2 binding partner in affinity purification and mass-spectrometry (AP-MS) experiments and I hypothesized that, like other tested ZFPs, ZNF121 is an RNA-binding protein and acts as a cofactor to stabilize YTHDF2 on shared mRNA targets. In this thesis, I utilize multi-omic and biochemical approaches to understand the role of ZNF121 in gene expression. I observe that ZNF121 is a cyto-nuclear protein and confirm its interaction with YTHDF2 and RNA. Using iCLIPseq, I found that ZNF121 binds mRNA around sites of YTHDF2 enrichment, even when no nearby m6A modification exists. Specifically, over 80% of ZNF121-bound transcripts are also YTHDF2-bound transcripts. Upon depletion of ZNF121, the binding of YTHDF2 to shared transcripts is impaired, and the stability of these transcripts increases in an m6A-independent fashion. This affects many transcripts coding for cell-cycle related proteins like MDM2, with effects on the DNA damage response and cell viability. Lastly, I observe that ZNF121 regulates cassette exon retention in the nucleus. My results show that ZNF121, beyond its conventional role as a transcription factor, is a versatile protein that regulates both mRNA stability in conjunction with YTHDF2 and mRNA splicing."]},{"key":"dc:description.degree","label":"Dc Description Degree","values":["Ph.D."]},{"key":"dc:title","label":"Title","values":["Uncovering the Roles of Human C2H2-Zinc Finger Protein ZNF121 in Gene Regulation"]}]}],"canonical_facts":{"dc:contributor.advisor":["Greenblatt, Jack F"],"dc:contributor.department":["Molecular Genetics"],"dc:creator":["Burke, Giovanni Livingston"],"dc:date":["2025-10"],"dc:date.accessioned":["2025-12-01T16:13:48Z"],"dc:date.issued":["2025-10"],"dc:description.abstract":["Gene expression is a complex, dynamic, and tightly orchestrated process. Regulation of genes transcribed by RNA polymerase II is coordinated by an elegant arrangement of protein machinery and chromatin architecture, and the resulting pre-mRNA is post-transcriptionally processed and regulated at every step to buffer, reduce, or amplify its protein output. Chemical nucleotide modifications on DNA or RNA are one type of gene regulatory mechanism. N6-methyladenosine (m6A) is the most abundant and well-characterized mRNA modification, facilitating mRNA regulatory processes that include mRNA decay regulated by the m6A reader proteins YTH-domain-containing family protein 2 (YTHDF2). However, YTHDF2 has a relatively weak affinity for its mRNA targets. RNA-binding proteins (RBPs) bind transcripts to exert regulatory effects, and many DNA-binding C2H2-zinc finger proteins (ZFPs) have recently been shown to be RBPs with roles in mRNA biology. I identified C2H2-ZFP, ZNF121, as a YTHDF2 binding partner in affinity purification and mass-spectrometry (AP-MS) experiments and I hypothesized that, like other tested ZFPs, ZNF121 is an RNA-binding protein and acts as a cofactor to stabilize YTHDF2 on shared mRNA targets. In this thesis, I utilize multi-omic and biochemical approaches to understand the role of ZNF121 in gene expression. I observe that ZNF121 is a cyto-nuclear protein and confirm its interaction with YTHDF2 and RNA. Using iCLIPseq, I found that ZNF121 binds mRNA around sites of YTHDF2 enrichment, even when no nearby m6A modification exists. Specifically, over 80% of ZNF121-bound transcripts are also YTHDF2-bound transcripts. Upon depletion of ZNF121, the binding of YTHDF2 to shared transcripts is impaired, and the stability of these transcripts increases in an m6A-independent fashion. This affects many transcripts coding for cell-cycle related proteins like MDM2, with effects on the DNA damage response and cell viability. Lastly, I observe that ZNF121 regulates cassette exon retention in the nucleus. My results show that ZNF121, beyond its conventional role as a transcription factor, is a versatile protein that regulates both mRNA stability in conjunction with YTHDF2 and mRNA splicing."],"dc:description.degree":["Ph.D."],"dc:identifier.uri":["https://hdl.handle.net/1807/150378"],"dc:rights":["Attribution 4.0 International"],"dc:rights.uri":["http://creativecommons.org/licenses/by/4.0/"],"dc:subject":["C2H2 Zinc Finger Protein","m6A","RNA biology","RNA modifications","YTHDF2","ZNF121"],"dc:title":["Uncovering the Roles of Human C2H2-Zinc Finger Protein ZNF121 in Gene Regulation"],"dc:type":["Thesis"]},"updated_at":"2026-07-27T21:28:20Z"}