MKRN3 imprinting, regulation, and action in the control of puberty

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ABSTRACT Puberty and reproduction are controlled by the hypothalamic-pituitary-gonadal (HPG) axis. The HPG axis is active during the embryonic and neonatal stages of human life but then suppressed during childhood. The re- activation of HPG axis results in puberty initiation. The precise mechanisms that regulate GnRH secretion to constrain the HPG axis during infancy and childhood and subsequently trigger the initiation of puberty remain elusive. The timing of puberty is associated with risks of subsequent disease and it is crucial to identify what elicits puberty initiation. Complex interactions among genetic, nutritional, and environmental factors influence pubertal initiation. We have recently identified loss-of-function mutations in MKRN3 in families with central precocious puberty (CPP). MKRN3 is located on chromosome 15q and is maternally imprinted, expressed only from the paternally inherited allele. The association of genetic mutations in MKRN3 with CPP is indisputable; however, the possibility of imprinting abnormalities in MKRN3 as a cause of pubertal disorders has not been explored. Epigenetic regulation of MKRN3 may be a link between environmental cues and pubertal timing. This proposal will investigate the MKRN3 imprinting profile during pubertal development and determine if MKRN3 imprinting abnormalities are associated with pubertal disorders; and study the molecular mechanisms by which MKRN3 regulates GnRH secretion. To this end, four distinct but complementary specific aims are proposed. The mentored phase of the proposal will be carried out under Dr. Ursula Kaiser's supervision at Brigham and Women's Hospital/Harvard Medical School. The aims of the K99 mentored phase are to: 1) Investigate the MKRN3 methylation profile in different phases of human life and confirm the methylation pattern in mouse tissues; and 2) Characterize the cellular and molecular mechanisms by which MKRN3 regulates GnRH secretion in vitro. This training will provide expertise in DNA methylation studies, in hiPSC experiments, and in RNA-Seq data analysis. The elucidation of the MKRN3 imprinting profile during normal pubertal development and an understanding of the actions of MKRN3 in hypothalamic neurons will provide a strong base of knowledge for the transition to the independent R00 phase of the award. Building on previous experience, the aims of the R00 independent phase are to: 3) Investigate if abnormalities in MKRN3 imprinting are associated with pubertal disorders; 4) Extend investigation of mechanisms of action of MKRN3 to in vivo mouse models. This award includes a well-structured training program that includes course work and seminar learning experiences. Completion of this project will lead to a better understanding of the molecular roles of MKRN3 in the regulation of GnRH secretion and advance our fundamental knowledge of MKRN3 imprinting. The successful completion of the aims of this proposal will bring new insights in the neuroendocrine regulation of GnRH secretion and enable me to establish my career as a successful independent translational investigator.