Sex-specific regulation of weight and puberty by the Lin28/let-7 axis

Abstract

Growth and pubertal timing differ in boys and girls. Variants in/near LIN28B associate with age at menarche (AAM) in genome-wide association studies and some AAM-related variants associate with growth in a sex-specific manner. Sex-specific growth patterns in response to Lin28b perturbation have been detected in mice, and overexpression of Lin28a has been shown to alter pubertal timing in female mice. To investigate further how Lin28a and Lin28b affect growth and puberty in both males and females, we evaluated Lin28b loss-of-function (LOF) mice and Lin28a gain-of-function (GOF) mice. Because both Lin28a and Lin28b can act via the conserved microRNA let-7, we also examined let-7 GOF mice. As reported previously, Lin28b LOF led to lighter body weights only in male mice while Lin28a GOF yielded heavier mice of both sexes. Let-7 GOF mice weighed less than controls, and males were more affected than females. Timing of puberty was assessed by vaginal opening (VO) and preputial separation (PS). Male Lin28b LOF and male let-7 GOF, but not female, mice displayed alteration of pubertal timing, with later PS than controls. In contrast, both male and female Lin28a GOF mice displayed late onset of puberty. Together, these data point toward a complex system of regulation by Lin28a, Lin28b, and let-7, in which Lin28b and let-7 can impact both puberty and growth in a sex-specific manner, raising the possibility that this pathway may contribute to differential regulation of male and female growth and puberty in humans.

Keywords: Lin28; let-7; puberty; sex-specific.

Figure 1

Growth patterns in Lin28a GOF (A–B), Lin28b LOF (C–D), and let-7 GOF (E–F) mice. The number of animals per group is indicated in each chart. Error bars represent SD. Growth data were analyzed with the student’s t-test at each time point. *P < 0.05.

Figure 2

The onset of puberty, as assessed by vaginal opening, VO, and preputial separation, PS, in Lin28a GOF, Lin28b LOF, and let-7 GOF mice. In each chart black boxes represent the GOF or LOF and open circles represent the control group. The inserted bar graphs display the mean day of VO or PS. Error bars represent SD. N=9–33, exact numbers per group are indicated in table 2. Puberty data were analyzed with the Mann-Whitney non-parametric t-test. *p<0.05.

Figure 3

Weight and age at puberty for each control (open circles) and transgenic (black boxes) mouse. The line represents linear regression of weight and age at VO/PS for controls. Significant differences in the relationship of age and weight at puberty between controls and transgenic mice were detected in both male and female Lin28a GOF mice and among let-7 GOF females.

Figure 4

Body fat content and glucose metabolism are similar in let-7 GOF mice and controls.(A–B) Body fat in 22-day-old let-7 GOF and littermate control mice, n=4–8; similar results were obtained for percent body fat (data not shown). (C–D) Intraperitoneal glucose tolerance test (IP-GTT) in 21-day-old let-7 GOF and littermate control mice, n=4–8. Error bars represent SD. Data were analyzed with the student’s t-test, assuming unequal variances. No data points reached P <0.05.

Figure 5

Hypothalamic let-7 expression levels were unaltered in Lin28b LOF and in let-7 GOF mice compared to controls. A. let-7a and let-7g levels in hypothalamus on the mean day of PS (postnatal day 27) or VO (postnatal day 30) among control Lin28b LOF mice, n=3–4. B let-7g levels in hypothalamus on the mean day of PS (postnatal day 26) or VO (postnatal day 29) among controls and let-7 GOF mice, n=3–4. Error bars represent SD. Expression data were analyzed using the student’s t-test. No data points reached P < 0.05.

Figure 6

Peripheral let-7 expression levels were untaltered in Lin28b LOF mice, but upregulated in in let-7 GOF mice. A–B. let-7a and let-7g levels in pituitary, gonads, and liver in Lin28b LOF mice on the mean day of or VO (postnatal day 30) or PS (postnatal day 27) among control mice, n=3–4. C–D. let-7g levels in pituitary, gonads, and liver on the mean day of VO (postnatal day 29) or PS (postnatal day 26) in let-7 GOF mice, n=3–4. Error bars represent SD. Expression data were analyzed using the student’s t-test. *P < 0.05.