Identification and characterization of two endoplasmic reticulum protein isoforms encoded by senescence-associated FAM134B gene

buir.advisorÖztürk, Mehmet
dc.contributor.authorTaşdemir, Nilgün
dc.date.accessioned2016-01-08T18:06:10Z
dc.date.available2016-01-08T18:06:10Z
dc.date.issued2008
dc.descriptionAnkara : The Department of Molecular Biology and Genetics and Institute of Engineering and Sciences of Bilkent University, 2008.en_US
dc.descriptionThesis (Master's) -- Bilkent University, 2008.en_US
dc.descriptionIncludes bibliographical refences.en_US
dc.description.abstractLiver cancer is the fifth most common cancer in the world. Until recently, tumor cells were known to have the capacity to proliferate indefinitely. In a previous study, we showed the spontaneous induction of replicative senescence in p53- and p16INK4a-deficient HCC (hepatocellular carcinoma) cells. In a follow-up study, we have analyzed the Affymetrix expression profiling of the senescent and immortal HCC clones that we had established. Among the genes with differential expression pattern, in this study, we have focused on a novel gene, FAM134B (family with sequence similarity 134, member B), which is significantly up-regulated (p-value=1.097E-06) in our senescent clones with respect to their immortal counterparts. FAM134B gene is located on human chromosome 5p15.1 near a LOH region, and its protein product has not yet been characterized. To begin with, we confirmed the up-regulation of FAM134B in our senescent clones as compared to our immortal clones by RT-PCR analysis. As a next step, meta-analysis of HCC microarray data indicated that the expression of FAM134B gene is progressively down-regulated in non-metastatic and metastatic HCC as compared to normal liver. Thus, we decided to characterize the protein product of this gene. Two known forms of transcripts were used to construct FLAG-tagged expression plasmids (encoding two isoforms with predicted molecular weights of 30 and 55 kDa). Immuno-staining experiments performed after transient ectopic expression indicated that both short and long isoforms of FAM134B-encoded protein localize to the endoplasmic reticulum (ER). Both protein isoforms co-localized with calnexin, a well known ER-chaperon. Thus, it appears that senescent cells over-express FAM134B-encoded ER protein isoforms, while cancer cells are deficient in their expression. We have also performed gain-of-function studies by stable ectopic expression of these two protein isoforms in an HCC cell line and addressed the potential role(s) of these isoforms in senescence and ER-stress. Our studies indicated that over-expression of these proteins did not have a ‘causative’ role in induction of senescence and did not affect the rate of cell proliferation. We also did not observe any changes in the responses of cells over-expressing these two protein isoforms to ER-stress induced via tunicamycin treatment. Therefore, FAM134B gene may be performing a yet unidentified function in senescent cells. All in all, we have identified two FAM134B-encoded proteins that localize to the ER, the function and the senescence association of which need further investigation.en_US
dc.description.provenanceMade available in DSpace on 2016-01-08T18:06:10Z (GMT). No. of bitstreams: 1 0003586.pdf: 10217877 bytes, checksum: e5ca30164b152ff1e2b4e8d7d0b8520e (MD5)en
dc.description.statementofresponsibilityTaşdemir, Nilgünen_US
dc.format.extentxviii, 112 leaves, illustrations, graphsen_US
dc.identifier.urihttp://hdl.handle.net/11693/14727
dc.language.isoEnglishen_US
dc.rightsinfo:eu-repo/semantics/openAccessen_US
dc.subject.lccQP624.75.P74 T37 2008en_US
dc.subject.lcshDNA-Protein interaction.en_US
dc.subject.lcshProtein interaction.en_US
dc.subject.lcshCancer cells.en_US
dc.subject.lcshp53 antioncogene.en_US
dc.subject.lcshLiver--Cancer.en_US
dc.subject.lcshEndoplasmic reticulum.en_US
dc.titleIdentification and characterization of two endoplasmic reticulum protein isoforms encoded by senescence-associated FAM134B geneen_US
dc.typeThesisen_US
thesis.degree.disciplineMolecular Biology and Genetics
thesis.degree.grantorBilkent University
thesis.degree.levelMaster's
thesis.degree.nameMS (Master of Science)

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