The bHLH-PAS transcription factor SIM1 is expressed during the development of the hypothalamic-pituitary axis in three hypothalamic nuclei: the paraventricular nucleus (PVN), the anterior periventricular nucleus (aPV), and the supraoptic nucleus (SON). To investigate Sim1 function in the hypothalamus, we produced mice carrying a null allele of Sim1 by gene targeting. Homozygous mutant mice die shortly after birth. Histological analysis shows that the PVN and the SON of these mice are hypocellular. At least five distinct types of secretory neurons, identified by the expression of oxytocin, vasopressin, thyrotropin-releasing hormone, corticotropin-releasing hormone, and somatostatin, are absent in the mutant PVN, aPV, and SON. Moreover, we show that SIM1 controls the development of these secretory neurons at the final stages of their differentiation. A subset of these neuronal lineages in the PVN/SON are also missing in mice bearing a mutation in the POU transcription factor BRN2. We provide evidence that, during development of the Sim1 mutant hypothalamus, the prospective PVN/SON region fails to express Brn2. Our results strongly indicate that SIM1 functions upstream to maintain Brn2 expression, which in turn directs the terminal differentiation of specific neuroendocrine lineages within the PVN/SON.[Key Words: SIM; hypothalamus; paraventricular nucleus; supraoptic nucleus; hormones]Received June 10, 1998; revised version accepted August 21, 1998.The hypothalamus integrates a variety of signals from the brain and the periphery to modulate secretion by the pituitary of peptidic hormones involved in maintaining homeostasis. A large body of work, starting with the discovery of oxytocin and vasopressin (Du Vigneaud 1955), has led to the identification of numerous neuropeptides secreted by hypothalamic and pituitary cells. The availability of these markers has allowed a detailed characterization of the different cell types found in the hypothalamus and the pituitary and the delineation of their physiological role. This fundamental work has made the hypothalamic-pituitary axis an advantageous system to study the mechanisms involved in generating cell diversity in the central nervous system (CNS).The neuroendocrine system consists of two sets of hypothalamic neurons: the magnocellular and the parvocellular neurons (for review, see Swanson and Sawchenko 1983;Swanson 1986;Sawchenko et al. 1992). The magnocellular neurosecretory system projects to the posterior pituitary where it releases vasopressin (AVP) and oxytocin (OT) directly into the general circulation. Vasopressin participates in the control of blood volume, osmolality, and pressure, whereas OT promotes parturition and lactation. The magnocellular neurons are located in two nuclei of the anterior hypothalamus, the paraventricular (PVN) and the supraoptic (SON) nuclei. Within the PVN and the SON, AVP and OT are produced by mutually exclusive sets of neurons. The sum of AVPand OT-producing cells corresponds to the total number of magnocellular neurons, indicating t...

One major function of the hypothalamus is to maintain homeostasis by modulating the secretion of pituitary hormones. The paraventricular (PVN) and supraoptic (SON) nuclei are major integration centers for the output of the hypothalamus to the pituitary. The bHLH-PAS transcription factor SIM1 is crucial for the development of several neuroendocrine lineages within the PVN and SON. bHLH-PAS proteins require heterodimerization for their function. ARNT, ARNT2, and BMAL1 are the three known general heterodimerization partners for bHLH-PAS proteins. Here, we provide evidence that Sim1 and Arnt2 form dimers in vitro, that they are co-expressed in the PVN and SON, and that their loss of function affects the development of the same sets of neuroendocrine cell types within the PVN and SON. Together, these results implicate ARNT2 as the in vivo dimerization partner of SIM1 in controlling the development of these neuroendocrine lineages.

It is important that chromosomes are duplicated only once per cell cycle. Over-replication is prevented by multiple mechanisms that block the reformation of a pre-replicative complex (pre-RC) onto origins in S and G2 phase. We have investigated the developmental regulation of Double-parked (Dup) protein, the Drosophila ortholog of Cdt1, a conserved and essential pre-RC component found in human and other organisms. We find that phosphorylation and degradation of Dup protein at G1/S requires cyclin E/CDK2. The N terminus of Dup, which contains ten potential CDK phosphorylation sites, is necessary and sufficient for Dup degradation during S phase of mitotic cycles and endocycles. Mutation of these ten phosphorylation sites, however, only partially stabilizes the protein,suggesting that multiple mechanisms ensure Dup degradation. This regulation is important because increased Dup protein is sufficient to induce profound rereplication and death of developing cells. Mis-expression has different effects on genomic replication than on developmental amplification from chorion origins. The C terminus alone has no effect on genomic replication,but it is better than full-length protein at stimulating amplification. Mutation of the Dup CDK sites increases genomic re-replication, but is dominant negative for amplification. These two results suggest that phosphorylation regulates Dup activity differently during these developmentally specific types of DNA replication. Moreover, the ability of the CDK site mutant to rapidly inhibit BrdU incorporation suggests that Dup is required for fork elongation during amplification. In the context of findings from human and other cells, our results indicate that stringent regulation of Dup protein is critical to protect genome integrity.