Hypothalamic neurodegeneration and adult-onset obesity in mice lacking the Ubb polyubiquitin gene - PubMed (original) (raw)

Hypothalamic neurodegeneration and adult-onset obesity in mice lacking the Ubb polyubiquitin gene

Kwon-Yul Ryu et al. Proc Natl Acad Sci U S A. 2008.

Abstract

Nearly all neurodegenerative diseases are associated with abnormal accumulation of ubiquitin (Ub) conjugates within neuronal inclusion bodies. To directly test the hypothesis that depletion of cellular Ub is sufficient to cause neurodegeneration, we have disrupted Ubb, one of four genes that supply Ub in the mouse. Here, we report that loss of Ubb led to a progressive degenerative disorder affecting neurons within the arcuate nucleus of the hypothalamus. This neurodegenerative cytopathology was accompanied by impaired hypothalamic control of energy balance and adult-onset obesity. Ubb was highly expressed in vulnerable hypothalamic neurons and total Ub levels were selectively reduced in the hypothalamus of Ubb-null mice. These findings demonstrate that maintenance of adequate supplies of cellular Ub is essential for neuronal survival and establish that decreased Ub availability is sufficient to cause neuronal dysfunction and death.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Fig. 1.

Fig. 1.

Abnormal growth of _Ubb_−/− mice. (A) Newborn _Ubb_−/− (−/−) (n = 18) mice are smaller and exhibit reduced perinatal weight gain compared with littermate controls (+/+ and +/−) (n = 39). Smaller size of _Ubb_−/− mice is also evident at E18.5 (+/+ and +/−, n = 13; −/−, n = 3). (B) Growth curve of male and female mice after weaning. Body weights of _Ubb_−/− (male, n = 19; female, n = 17) mice are significantly lower than those of their littermate controls (male, n = 51; female, n = 76) before 16 weeks of age. At 16 weeks of age, there is no significant difference in weight between _Ubb_−/− mice and littermate controls. (C) Total body mass of male mice determined by DEXA (+/+ and +/−, n = 4–12; −/−, n = 3 or 4). (D) Snout–anus length of 4-month-old mice (+/+ and +/−, n = 18; −/−, n = 13). (E) Representative photographs of adult Ubb+/+ (+/+) and _Ubb_−/− littermates. All data are expressed as means ± SEM from the indicated number of mice. ∗, P < 0.05; ∗∗, P < 0.01; ∗∗∗, P < 0.001 for _Ubb_−/− vs. littermate controls.

Fig. 2.

Fig. 2.

Altered body composition in adult _Ubb_−/− mice. (A) Whole-body fat content of male _Ubb_−/− mice (n = 3 or 4) and littermate controls (n = 4–12) measured by DEXA. Fat content is expressed as a percentage of body weight. (B) Inguinal fat pad weight of male (+/+, n = 6–11; −/−, n = 6–11) and female (+/+, n = 5 or 7; −/−, n = 6) mice. (C) Fat and lean mass of male _Ubb_−/− mice (n = 3 or 4) and littermate controls (n = 4–12) measured by DEXA. All data are expressed as means ± SEM from the indicated number of mice. ∗, P < 0.05; ∗∗, P < 0.01; ∗∗∗, P < 0.001 for _Ubb_−/− vs. littermate controls or Ubb+/+.

Fig. 3.

Fig. 3.

Defective central regulation of body weight and energy homeostasis in _Ubb_−/− mice. (A) Ad libitum food intake during the 24 h preceding fasting for 48 h and the two 24-h intervals after refeeding was measured and expressed as per gram body weight in 7-week-old (+/+, n = 9; −/−, n = 11) and 15-week-old (+/+, n = 11; −/−, n = 11) mice. (B) Recovery of body weight after fasting for 48 h and refeeding for 48 h in 7-week-old (+/+, n = 9; −/−, n = 11) and 15-week-old (+/+, n = 11; −/−, n = 11) mice. Loss of body weight after fasting was arbitrarily assigned as −100% in each genotype of mice. (C) Abnormal expression of hypothalamic neuropeptides in adult _Ubb_−/− mice. Normalized mRNA levels, measured by quantitative real-time RT-PCR from hypothalamus of young (5–6 weeks old) and adult (3–4 months old) ad libitum fed (+/+ and +/−, n = 12; −/−, n = 5 or 6) or 48-h fasted (+/+ and +/−, n = 12 or 16; −/−, n = 6) mice. All data are expressed as means ± SEM from the indicated number of mice. In A and C: ∗, P < 0.05; ∗∗, P < 0.01; #, P = 0.07. In B: ∗∗∗, P < 0.001 vs. 15-week-old Ubb+/+ mice after refeeding for 48 h.

Fig. 4.

Fig. 4.

Degeneration of hypothalamic neurons in adult _Ubb_−/− mice. In situ hybridization of AgRP (A) and NSE (B) mRNA from _Ubb_−/− mice (n = 3 or 4) and littermate controls (n = 3 or 4). Images are representative for each age and group. For quantification, stereological counts were obtained from eight serial sections generated from each brain. 3V, third ventricle. The data are expressed as means ± SEM from the indicated number of mice. ∗, P < 0.05.

Fig. 5.

Fig. 5.

Dominant contribution of Ubb to Ub content of hypothalamus. (A) Quantification of ubiquitin transcript levels in hypothalamus of 3- to 4-month-old mice. (Left) Indicated mRNA levels, measured by quantitative real-time RT-PCR (+/+, n = 5; +/−, n = 6; −/−, n = 4). (Right) Relative contribution of ubiquitin genes to total Ub-coding potential normalized to the number of Ub moieties encoded by each ubiquitin transcript. The data are expressed as means ± SEM from the indicated number of mice. (B) Total Ub content in whole brain (n = 6 per genotype) and hypothalamus (+/+, n = 6; +/−, n = 3; −/−, n = 4) from 5- to 6-month-old mice. The data are expressed as means ± SEM from the indicated number of mice. ∗∗∗, P < 0.001 vs. Ubb+/+ mice. (C) Neuronal pattern of Ubb expression in the arcuate nucleus. Representative confocal images of GFP-Puror and Nissl stain from 4-month-old _Ubb_−/− mice (n = 3) are shown. The far right image shows the arcuate nucleus at higher magnification. VMH, ventromedial hypothalamus; ARC, arcuate nucleus; 3V, third ventricle. (Scale bars, 100 μm.) (D) Colocalization of NPY, AgRP, and α-MSH immunoreactive neurons in the arcuate nucleus to _Ubb_-expressing neurons. Representative confocal images of GFP-Puror and NPY, AgRP, and α-MSH immunoreactivity from 4-month-old Ubb+/− mice (n = 4) are shown. 3V, third ventricle. (Scale bars, 100 μm.)

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