Year: 2013
Type: dissertation
Abstract: SUMOylation is an evolutionary conserved post-translational regulatory mechanism involving the covalent attachment of a Small Ubiquitin-like MOdifier (SUMO) to target proteins in reactions catalyzed by E1, E2 and E3 enzymes. The consequences of protein SUMOylation are varied; it can affect protein localization, create new interaction surfaces, exclude others, regulate transcription factor activity, and protect proteins from destruction. Loss of SUMOylation is not compatible with life, as knock-out of the only E2 enzyme (Ubc9) leads to embryonal lethality in mice. SUMOylation is possible but less specific and efficient in the absence of E3 enzymes, of which the protein inhibitors of activated STAT (PIAS) family is the most widely studied. In mammals, there are three conjugatable SUMO paralogs of which SUMO-1 differs from SUMO2/3 the most and lacks the ability to form (poly)SUMO chains. SUMO2/3 are encoded by two separate genes, but differ only by three amino acids leading to many overlapping functions. The purpose of this thesis was to study the in vivo functions of SUMOylation by using knock-out animal models. PIAS2 is an E3 enzyme highly expressed in testis and involved in androgen signaling. A Pias2 null mouse line was generated and found to be fertile and viable. Loss of Pias2 led to a decrease in testicular weight and sperm count and increase in apoptotic cells in the seminiferous tubules. However, the sperm was qualitatively normal, potentially due to compensation by other PIAS proteins. When Sumo1 was knocked out by targeted disruption, the mice were born in normal Mendelian ratios, and were viable and fertile. At least in the case of Ran GTPase-activating protein 1 (RanGAP1), the most heavily SUMO-1 modified protein, lack of SUMO-1 resulted in its SUMOylation by SUMO-2/3. This compensatory mechanism, however, was not reflected in the mRNA levels. In testis, SUMO-1 was localized to a nuclear structure called sex vesicle present in pachytene spermatocytes. SUMO-2/3 was also present there, and the structure was able to form in the absence of SUMO-1. The phenotypes of the Sumo1 null and wild-type mice did not differ under normal laboratory conditions. However, when the mice were challenged with lipopolysaccharide (LPS), a bacterial antigen, Sumo1 null mice developed a more pronounced acute phase response in liver due to inefficient transrepressive function of liver receptor homolog 1 (LRH-1) leading to more efficient removal of repressive factors from inflammatory gene promoters. Another nuclear receptor, liver X receptor (LXR), required SUMO-2/3 modification to function properly testifying that SUMO paralogs can partly but not in all cases compensate for each other. Another example found in this study where Sumo1 function cannot be compensated for was adipogenesis. Adipogenesis is highly dependent on the functions of peroxisome proliferator-activated receptor (PPAR) γ. It was found that, contrary to previous reports, gene activation functions of PPARγ are compromised in the absence of SUMO-1 as shown by inefficient transactivation of a luciferase reporter in Sumo1 null cells and incomplete response of Sumo1 knock-out mice to rosiglitazone, a PPARγ agonist. Furthermore, embryonic fibroblasts and mesenchymal stem cells of Sumo1 null mice and SUMO-1-depleted preadipocytes differentiated less efficiently into adipocytes, further demonstrating the importance of SUMO-1 in the activation functions of PPARγ. When Sumo1 null mice were challenged with a high-fat diet, they gained less weight than their wild-type littermates due to a limited expansion of white adipose tissue. The results show that SUMO-1 is important for the activation functions of PPARγ either via modification of PPARγ or a coregulatory protein. Taken together, the results of this thesis show that the SUMOylation pathway has many redundant functions. However, this study identified physiological phenomena where the lack of Pias2 or Sumo1 could not be compensated in vivo.
Source: Työväentutkimus Vuosikirja
Author: Laura Mikkonen
Language: English
Subfield: Molecular Biology
Domain: Life Sciences
Open Access status: green
OpenAlex ID: W1551974835