Uloga proteina BPM1 u mehanizmu metilacije DNA de novo tijekom razvoja uročnjaka (Arabidopsis thaliana L.)

doctoral thesis

• Title (english): The role of BPM1 protein in de novo DNA methylation mechanism during development of Arabidopsis thaliana L.
• Author: Tamara Vuk
• Mentor: Dunja Leljak-Levanić (mentor)
• Committee member: Mirjana Pavlica (predsjednik povjerenstva)
• Committee member: Biljana Balen (član povjerenstva)
• Committee member: Snježana Mihaljević (član povjerenstva)
• Granter: University of Zagreb; Faculty of Science; (Department of Biology); Zagreb
• Defense date and country: 2023-01-24, Croatia
• Scientific / art field, discipline and subdiscipline: NATURAL SCIENCES; Biology
• Universal decimal classification (UDC): 577 - Biochemistry. Molecular biology. Biophysics

Abstract

Proteini BPM iz vrste Arabidopsis thaliana primarnu ulogu ostvaruju u putu
proteasomalne razgradnje ovisne o ubikvitinu, u sklopu kojeg prepoznaju specifične supstrate
za razgradnju. Prethodno je utvrđena interakcija proteina BPM1 s proteinima RDM1 i DMS3,
komponentama metilacijskog mehanizma RdDM. Uloga proteina BPM1 u mehanizmu RdDM
u ovom je radu ispitana primjenom kromatinske imunoprecipitacije u svrhu identifikacije novih
ciljnih gena ovog mehanizma u čijoj regulaciji metilacije posreduje protein BPM1.
Identificirani su geni FBW2, RKP i AGL14, a kao referentni odabran je gen CML41. Analizom
metilacije ovih gena metodom bisulfitnog sekvenciranja i pirosekvenciranja u linijama s
prekomjernom ekspresijom proteina od interesa (oeBPM1, oeDMS3), te linijama s narušenom
funkcijom mehanizma RdDM (dms3-1, nrpe1-10), potvrđen je stimulativni učinak proteina
BPM1 na aktivnost mehanizma, koji je najizraženiji tijekom embriogeneze, a sam mehanizam
RdDM značajniji je za zigotnu nego za somatsku embrigenezu. Ovisno o poziciji, učinci
metilacijskih događaja reflektiraju se na ekspresiju svih analiziranih gena.

Abstract (english)

Proteins BPM of Arabidopsis thaliana have a primary role in the ubiquitin-dependent
proteasomal degradation pathway, where they recognize specific substrates for degradation.
The interaction of BPM1 protein with RDM1 and DMS3, components of RdDM methylation
mechanism, has previously been established. Here, the role of BPM1 protein in RdDM was
analyzed using chromatin immunoprecipitation for identification of new RdDM targets whose
methylation is mediated by BPM1 protein. Genes FBW2, RKP and AGL14 were identified as
targets, and CML41 was chosen as a reference gene. Methylation analysis of these genes was
conducted using bisulfite sequencing and pyrosequencing in lines with overexpression of
BPM1 (oeBPM1) or DMS3 (oeDMS3), and lines with impaired RdDM function (dms3-1,
nrpe1-10). The results confirmed a stimulative role of BPM1 protein in RdDM. This
mechanism proved to be more significant in zygotic than somatic embyrogenesis. Depending
on the position, methylation had a diverse effect on expression of analyzed genes.

• Keywords: Arabidopsis thaliana; BPM1; metilacija; RdDM; ChIP; pirosekvenciranje
• Keywords (english): Arabidopsis thaliana; BPM1; methylation; RdDM; ChIP; pyrosequencing
• Language: croatian
• URN:NBN: urn:nbn:hr:217:895020
• Promotion: 2023
• Study programme: Title: Biology; Study programme type: university; Study level: postgraduate; Academic / professional title: doktor/doktorica znanosti, područje prirodnih znanosti, polje biologija (doktor/doktorica znanosti, područje prirodnih znanosti, polje biologija)
• Type of resource: Text
• File origin: Born digital
• Access conditions: Embargoed access; Embargo expiration date: 2025-01-24
• Created on: 2023-01-31 12:20:09