Award Date

5-1-2021

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Department

Life Sciences

First Committee Member

Nora Caberoy

Second Committee Member

Allen Gibbs

Third Committee Member

Frank Van Breukelen

Fourth Committee Member

Jefferey Shen

Fifth Committee Member

Jefferson Kinney

Number of Pages

133

Abstract

A homozygous mutation in tub gene causes retinal and cochlear degeneration and adult-onset obesity in mice, and retinal dystrophy and obesity in humans. A combination of these phenotypes is seen in human syndromes with unknown molecular mechanisms. Thus, elucidating the pathways where Tubby protein participates in is crucial in understanding how these disease phenotypes developed and in formulating therapeutic strategies for the prevention or treatment of these diseases.

Tubby protein has been demonstrated to participate in both extra- and intracellular functions. In the retina, Tubby is involved in the clearance of dead photoreceptor cells. Tubby facilitates phagocytosis in the retina by acting as a bridge between the dead photoreceptor outer segments and the Mer tyrosine kinase (MerTK) receptor of the retinal pigment epithelium. Tubby also promotes microglial phagocytosis of apoptotic cells through the MerTK receptor. In addition to these, Tubby acts as an intracellular effector of G-protein coupled receptor (GPCR) signaling, and is involved in ciliary transport. However, these molecular processes are not enough to explain the pathology behind the three disease phenotypes that resulted from a mutation in tub gene. To gain a better understanding of the molecular functions of Tubby, our lab identified Tubby-interacting proteins and investigated how these interactions associate with tubby mutant disease phenotypes.

This study identified Estrogen related receptor beta (Esrrb) as a Tubby-interacting protein. The interaction occurs in vivo and is mediated by the N-terminal domain of Tubby and DNA-binding domain of Esrrb. Furthermore, we have demonstrated that Esrrb is a direct transcriptional target of Tubby protein. Tubby translocates into the nucleus, binds to chromatin, associates with the basal transcription machinery and other chromatin-interacting proteins, and transcriptionally activates Esrrb by binding to the ERRE in the Esrrb promoter. Upregulation of Esrrb inhibits transcriptional activation by Tubby, suggesting a negative feedback loop where Esrrb inhibits its own expression. Loss of function mutation of Esrrb has been reported to result in reduced rod photoreceptor survival. Esrrb is downregulated in tubby mutant retina, which could explain the progressive photoreceptor loss observed. The results of this study demonstrate that Tubby is a transcription factor that regulates expression of Esrrb, thus providing a novel mechanistic insight on how Tubby promotes photoreceptor survival through Esrrb induction. As Esrrb is also a regulator of genes involved in maintaining energy balance in the retina, and tubby mutants exhibit defective carbohydrate metabolism and energy imbalance, Tubby may be regulating energy homeostasis through Esrrb.

This study further explored the role of Tubby in adult-onset obesity. The high expression of Tubby in neurons suggests its role in central regulation of energy homeostasis, which has been emphasized in previous studies. However, the role of Tubby in adipose tissues and peripheral regulation of energy balance remains unexplored. Previous findings have demonstrated the role of Hmgb2 in promoting differentiation of mesenchymal stem cells into adipocytes. We have previously identified Hmgb2 as a putative Tubby-binding protein. Thus, we investigated the possible role of Tubby-Hmgb2 interaction in adipogenesis and fat accumulation to gain insight into the mechanism of obesity development in tubby mutants. Here we show that Hmgb2 interacts with Tubby. Tubby and Hmgb2 are present in the cytoplasm and nucleus of preadipocytes. In addition, Hmgb2 is found in the chromatin bound fraction. We also demonstrated that overexpression of Tubby in 3T3-L1 adipocytes prevents fat accumulation. Subcutaneous white adipose tissues from tubby mice showed increased in Hmgb2 protein levels. In contrast, proadipogenesis factors such as C/EBPa, C/EBPb, and PPARg were decreased, which may indicate impaired adipogenesis. These data suggest that Tubby may regulate adipogenesis through its interaction with Hmgb2 and regulation of proadipogenesis factors. Taken together, these findings suggest that retinal degeneration and obesity phenotypes in tubby mutants could arise from disrupting the role of Tubby in maintaining energy homeostasis.

Keywords

Adipogenesis; Esrrb; Hmgb2; Transcription factor; Tubby

Disciplines

Biology | Cell Biology | Molecular Biology

File Format

pdf

File Size

4000 KB

Degree Grantor

University of Nevada, Las Vegas

Language

English

Rights

IN COPYRIGHT. For more information about this rights statement, please visit http://rightsstatements.org/vocab/InC/1.0/

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Available for download on Monday, May 15, 2028


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