A study of the expression and cellular function of the human FAM111B gene

dc.contributor.advisorArowolo, Afolake
dc.contributor.authorRhoda, Cenza
dc.date.accessioned2022-03-16T06:38:35Z
dc.date.available2022-03-16T06:38:35Z
dc.date.issued2021
dc.date.updated2022-03-16T00:53:38Z
dc.description.abstractPOIKTMP, a multi-systemic fibrosing disease, results from mutations in the human FAM111B gene. Studies have also suggested high expression of this gene in cancers. Despite rising interest in the pathological effects of FAM111B mutations and overexpression of FAM111B, knowledge of the physiological role of this gene remains limited. Therefore, this study sought out to provide insights into the cellular function of FAM111B and to investigate the pathological effect of the FAM111B Y621D mutation. First, bioinformatics studies coupled with quantitative PCR and Western blots analysis were employed to assess FAM111B gene and protein expression in cancerous and non-cancerous cell lines. Subsequently, FAM111B gene expression was downregulated and upregulated in the human fibrosarcoma (HT1080) cell line by RNA-interference mediated gene silencing and recombinant gene expression technologies. The effect of these FAM111B dysregulations was studied using cellbased functional assays: proliferation, apoptosis, migration, and invasion assays. Furthermore, the functional pathways and interacting proteins of the FAM111B protein was determined using mass spectroscopy proteomics. Finally, preliminary studies in a POIKTMP patient-derived fibroblasts were attempted to recapitulate the results obtained using the HT1080 cell line. The results from this study indicated that FAM111B gene and protein overexpression occurs in cancer cells. Second, the depletion of FAM111B suggests a decelerated rate of cell proliferation and migration (14%), and increased apoptosis (1.4-fold). Conversely, overexpression of FAM111B resulted in a marked reduction in apoptosis (3-fold) and increased cell migration by 27 %, howbeit, no evidence of increased proliferation. Furthermore, Y621D FAM111B mutant cells showed reduced expression of FAM111B, decreased apoptosis (1.1-fold), cellular invasion (24%), and indicates an increase in cell proliferation and migration (18 %). The proteomics data suggested wild-type FAM111B interacts with HSP7C, a molecular chaperone, which alongside BAG3 and BCL2 to minimise apoptosis. Similarly, Y621D's interaction with G3V3W4, a component of the 20S proteasome complex involved in the proteolytic degradation of damaged proteins, may suggest the rapid clearance of this mutant protein.
dc.identifier.apacitationRhoda, C. (2021). <i>A study of the expression and cellular function of the human FAM111B gene</i>. (). ,Faculty of Health Sciences ,Department of Medicine. Retrieved from http://hdl.handle.net/11427/36135en_ZA
dc.identifier.chicagocitationRhoda, Cenza. <i>"A study of the expression and cellular function of the human FAM111B gene."</i> ., ,Faculty of Health Sciences ,Department of Medicine, 2021. http://hdl.handle.net/11427/36135en_ZA
dc.identifier.citationRhoda, C. 2021. A study of the expression and cellular function of the human FAM111B gene. . ,Faculty of Health Sciences ,Department of Medicine. http://hdl.handle.net/11427/36135en_ZA
dc.identifier.ris TY - Master Thesis AU - Rhoda, Cenza AB - POIKTMP, a multi-systemic fibrosing disease, results from mutations in the human FAM111B gene. Studies have also suggested high expression of this gene in cancers. Despite rising interest in the pathological effects of FAM111B mutations and overexpression of FAM111B, knowledge of the physiological role of this gene remains limited. Therefore, this study sought out to provide insights into the cellular function of FAM111B and to investigate the pathological effect of the FAM111B Y621D mutation. First, bioinformatics studies coupled with quantitative PCR and Western blots analysis were employed to assess FAM111B gene and protein expression in cancerous and non-cancerous cell lines. Subsequently, FAM111B gene expression was downregulated and upregulated in the human fibrosarcoma (HT1080) cell line by RNA-interference mediated gene silencing and recombinant gene expression technologies. The effect of these FAM111B dysregulations was studied using cellbased functional assays: proliferation, apoptosis, migration, and invasion assays. Furthermore, the functional pathways and interacting proteins of the FAM111B protein was determined using mass spectroscopy proteomics. Finally, preliminary studies in a POIKTMP patient-derived fibroblasts were attempted to recapitulate the results obtained using the HT1080 cell line. The results from this study indicated that FAM111B gene and protein overexpression occurs in cancer cells. Second, the depletion of FAM111B suggests a decelerated rate of cell proliferation and migration (14%), and increased apoptosis (1.4-fold). Conversely, overexpression of FAM111B resulted in a marked reduction in apoptosis (3-fold) and increased cell migration by 27 %, howbeit, no evidence of increased proliferation. Furthermore, Y621D FAM111B mutant cells showed reduced expression of FAM111B, decreased apoptosis (1.1-fold), cellular invasion (24%), and indicates an increase in cell proliferation and migration (18 %). The proteomics data suggested wild-type FAM111B interacts with HSP7C, a molecular chaperone, which alongside BAG3 and BCL2 to minimise apoptosis. Similarly, Y621D's interaction with G3V3W4, a component of the 20S proteasome complex involved in the proteolytic degradation of damaged proteins, may suggest the rapid clearance of this mutant protein. DA - 2021_ DB - OpenUCT DP - University of Cape Town KW - Medicine LK - https://open.uct.ac.za PY - 2021 T1 - A study of the expression and cellular function of the human FAM111B gene TI - A study of the expression and cellular function of the human FAM111B gene UR - http://hdl.handle.net/11427/36135 ER - en_ZA
dc.identifier.urihttp://hdl.handle.net/11427/36135
dc.identifier.vancouvercitationRhoda C. A study of the expression and cellular function of the human FAM111B gene. []. ,Faculty of Health Sciences ,Department of Medicine, 2021 [cited yyyy month dd]. Available from: http://hdl.handle.net/11427/36135en_ZA
dc.language.rfc3066eng
dc.publisher.departmentDepartment of Medicine
dc.publisher.facultyFaculty of Health Sciences
dc.subjectMedicine
dc.titleA study of the expression and cellular function of the human FAM111B gene
dc.typeMaster Thesis
dc.type.qualificationlevelMasters
dc.type.qualificationlevelMSc
Files
Original bundle
Now showing 1 - 1 of 1
Loading...
Thumbnail Image
Name:
thesis_hsf_2021_rhoda cenza.pdf
Size:
2.99 MB
Format:
Adobe Portable Document Format
Description:
License bundle
Now showing 1 - 1 of 1
Loading...
Thumbnail Image
Name:
license.txt
Size:
0 B
Format:
Item-specific license agreed upon to submission
Description:
Collections