Brenda Gerhardt

Regulators of the Proteasome Pathway, Uch37 and Rpn13, Play Distinct Roles in Mouse Development

Rpn13 is a novel mammalian proteasomal receptor that has recently been identified as an amplification target in ovarian cancer. It can interact with ubiquitin and activate the deubiquitinating enzyme Uch37 at the 26S proteasome. Since neither Rpn13 nor Uch37 is an integral proteasomal subunit, we explored whether either protein is essential for mammalian development and survival. Deletion of Uch37 resulted in prenatal lethality in mice associated with severe defect in embryonic brain development. In contrast, the majority of Rpn13-deficient mice survived to adulthood, although they were smaller at birth and fewer in number than wild-type littermates. Absence of Rpn13 produced tissue-specific effects on proteasomal function: increased proteasome activity in adrenal gland and lymphoid organs, and decreased activity in testes and brain. Adult Rpn13−/− mice reached normal body weight but had increased body fat content and were infertile due to defective gametogenesis. Additionally, Rpn13−/− mice showed increased T-cell numbers, resembling growth hormone-mediated effects. Indeed, serum growth hormone and follicular stimulating hormone levels were significantly increased in Rpn13−/− mice, while growth hormone receptor expression was reduced in the testes. In conclusion, this is the first report characterizing the physiological roles of Uch37 and Rpn13 in murine development and implicating a non-ATPase proteasomal protein, Rpn13, in the process of gametogenesis.

Differential sensitivity to SSRI and tricyclic antidepressants in juvenile and adult mice of three strains.

Clinical studies have shown differential efficacy of several antidepressants in children and adolescents compared to adults, yet few animal studies have sought to characterize this phenomenon. We compared effects of fluoxetine and imipramine in two common behavioral assays that hold high predictive validity for antidepressant activity, tail suspension and forced swim test, using juvenile (5 weeks) and adult (12 weeks) mice from 3 strains. C57BL/6J-Tyr(c-Brd) (C57), hybrid C57BL/6J-Tyr(c-Brd)x129S5/SvEvBrd (F2), and Balb/cAnNTac (Balb/C) mice were tested in forced swim test and tail suspension after i.p. dosing with either fluoxetine or imipramine. Brain tissues were analyzed to evaluate levels of VMAT2, a possible modulator of age-dependent sensitivity to antidepressants. Imipramine had more consistent antidepressant effect across age groups and strains. Imipramine increased struggle in mice of both ages. Fluoxetine did not have an effect on immobility in Balb/C of both ages in tail suspension. Fluoxetine also did not increase forced swim struggle behavior in juvenile mice of all strains, but was effective in increasing struggle in adults. Juvenile mice had higher immobility and lower struggle than adults in forced swim, and juveniles also had higher immobility in tail suspension test for Balb/C and C57. In addition, VMAT2 levels were increased in juveniles. These results confirm that standard antidepressants produce effects in both juveniles and adults but age-related differences were evident in both tests. Further examination of these effects is needed to determine whether it may be related to age-dependent difference in the clinical response to antidepressants of these classes.

Profound Obesity Secondary to Hyperphagia in Mice Lacking Kinase Suppressor of Ras 2

The kinase suppressor of ras 2 (KSR2) gene resides at human chromosome 12q24, a region linked to obesity and type 2 diabetes (T2D). While knocking out and phenotypically screening mouse orthologs of thousands of druggable human genes, we found KSR2 knockout (KSR2−/−) mice to be more obese and glucose intolerant than melanocortin 4 receptor−/− (MC4R−/−) mice. The obesity and T2D of KSR2−/− mice resulted from hyperphagia which was unresponsive to leptin and did not originate downstream of MC4R. The kinases AMP‐activated protein kinase (AMPK) and mammalian target of rapamycin (mTOR) are each linked to food intake regulation, but only mTOR had increased activity in KSR2−/− mouse brain, and the ability of rapamycin to inhibit food intake in KSR2−/− mice further implicated mTOR in this process. The metabolic phenotype of KSR2 heterozygous (KSR2+/minus;) and KSR2−/− mice suggests that human KSR2 variants may contribute to a similar phenotype linked to human chromosome 12q24.