A proteomics-based approach to investigate proteins associated with adipogenesis

ResearchSpace Repository

Show simple item record

dc.contributor.advisor Cooper, Garth J.S en
dc.contributor.advisor Xu, Aimin en
dc.contributor.author Choi, Kai-Luk, 1966- en
dc.date.accessioned 2007-08-06T11:58:38Z en
dc.date.available 2007-08-06T11:58:38Z en
dc.date.issued 2004 en
dc.identifier.citation Thesis (PhD--Biological Sciences)--University of Auckland, 2004 en
dc.identifier.uri http://hdl.handle.net/2292/1297 en
dc.description Full text is available to authenticated members of The University of Auckland only. en
dc.description.abstract Adipose tissue has recently been shown to play an important role in the regulation of energy metabolism. Malfunction of adipose tissue is one of the major causes-of insulin resistance and its associated complications, such as type 2 diabetes and cardiovascular disease. Studies of adipocytes are therefore important. In this study, a differential proteome mapping strategy was used to identify intracellular proteins whose expression was substantially altered during conversion of mouse 3T3-L1 preadipocytes to adipocytes. Two-dimensional gel electrophoresis analysis identified ten proteins, which were induced following hormone-insulin evoked differentiation. Nine of the ten proteins were found to have up-regulated expression or were predominantly expressed in adipocytes, and one of these was a novel protein. One of the ten proteins was diminished and identified as an α2 macroglobulin-like protein fragment. The novel protein and the diminished protein were then further investigated in this study. The first protein chosen for further study was the novel protein. We called it Fat tissue-specific Low molecular weight Protein (FALP). It was preferentially expressed in adipocytes but not in preadipocytes. The falp cDNA was cloned both from mouse (accession number AY079153) and two isoforms from human (accession numbers AY078152 and AF4835) using degenerate PCR. Northern blot analysis showed that the falp gene was predominantly expressed in brown and white fat tissue. Human homologues of mouse FALP were found to exist as two alternatively spliced isoforms, which shared the same NH2-terminus but with different COOH-termini. A sequence homology search revealed that FALP did not share sequence identity with any genes of known function. Both human and mouse FALP contained a conserved putative transmembrane domain. Immunocytochemistry analyses suggested that FALP localized at mitochondria. It interacted with glutaryl CoA dehydrogenase as demonstrated by in vitro pull-down and in vivo co-immunoprecipitation. These data suggested FALP might be involved in fatty acid metabolism during adipogenesis. The second protein studied was the α2 macroglobulin-like protein fragment. It was found to be present in very high abundance in 3T3-L1 preadipocytes. It was dramatically decreased after hormone-insulin induced differentiation of preadipocytes to adipocytes. Analysis by metabolic radiolabelling with [35S] methionine indicated that the intracellular accumulation of the α2 macroglobulin fragment was derived from the extracellular medium and not by de novo synthesis. A similar phenomenon also occurred in primary preadipocytes (stromal-vascular cells) and adipocytes isolated from mouse peri-uterine fat pads. Incubation of 3T3-L1 preadipocytes with an anti-α2 macroglobulin neutralising antibody caused depletion of the intracellular α2 macroglobulin fragment, and also enhanced spontaneous adipocyte differentiation. These results suggest that: 1. Intracellular accumulation of the α2 macroglobulin fragment could inhibit adipocyte differentiation, and 2. Differentiation can be induced, at least in part by suppressing the activity of intracellular α2 macroglobulin in 3T3-L1 preadipocytes. In summary, the results of this work have identified two new regulatory mechanisms related to the conversion of preadipocytes into adipocytes, and the regulation of adipocyte metabolism. en
dc.language.iso en en
dc.publisher ResearchSpace@Auckland en
dc.relation.ispartof PhD Thesis - University of Auckland en
dc.relation.isreferencedby UoA99132224714002091 en
dc.rights Restricted Item. Available to authenticated members of The University of Auckland. en
dc.rights.uri https://researchspace.auckland.ac.nz/docs/uoa-docs/rights.htm en
dc.title A proteomics-based approach to investigate proteins associated with adipogenesis en
dc.type Thesis en
thesis.degree.discipline Biological Sciences en
thesis.degree.grantor The University of Auckland en
thesis.degree.level Doctoral en
thesis.degree.name PhD en
dc.rights.holder Copyright: The author en

Files in this item

Find Full text

This item appears in the following Collection(s)

Show simple item record


Search ResearchSpace

Advanced Search