Cloning and Functional Analysis of Hypothalamic Homeobox Gene Bsx1a and Its Isoform, Bsx1b

Abstract

The hypothalamus is a key regulatory unit of the neuroendocrine system and plays an essential role in energy balance and reproduction. Despite its important role, the molecular mechanisms underlying hypothalamic development are not fully understood. Here, we report molecular analyses of a newly identified murine homeobox gene, Bsx/Bsx1a, that is expressed in the developing and postnatal hypothalamus. We demonstrate that BSX1A is a DNA binding protein and a transcriptional activator. Transcriptional reporter assays identified the C-terminal region of BSX1A as an activation domain. We have isolated an alternative splice form of Bsx1a, designated Bsx1b, which retains the N-terminal region but lacks the homeodomain. Analyses of subcellular localization using transfected cell lines revealed that BSX1A and BSX1B localize in the nuclei and cytoplasm, respectively. Immunohistochemical analyses suggested that both BSX1A and BSX1B are expressed in the neonatal hypothalamus. Taking these data together, we propose that alternative RNA splicing is involved in hypothalamic development/function.The hypothalamus-pituitary axis is a major component of the neuroendocrine system that controls the homeostasis of energy balance and reproduction. Disturbances of hypothalamic functions cause a variety of human health problems that include growth retardation, obesity, infertility, and hormoneregulated tumor progression. Much has been elucidated in the development of the pituitary in terms of transcriptional regulation; however, relatively little is known about the development of hypothalamic neurons (4,5,16,28,31,35).Although the hypothalamus consists of several cell types located in different regions, only a few transcription factors have been identified that are expressed in the developing hypothalamus. Brn-2 encodes a POU domain transcription factor, and its targeted mutation results in the loss of corticotropin-releasing hormone (CRH), vasopressin (AVP), and oxytocin (OT) in Brn-2-expressing cells. The homozygous mutant mice die in the neonatal period with a decrease in size and weight (18,26). The basic helix-loop-helix (bHLH)-Per-ArntSim transcription factor Sim1 is expressed in the paraventricular, the supraoptic, and the anterior periventricular nuclei. In Sim1 homozygous mutant mice, OT-, AVP-, thyrotropin-releasing hormone-, CRH-, and somatostatin (SS)-secreting neurons are absent in the Sim1-expressing regions (15). The Sim1 mutant mice die within 24 h after birth. Thus, the phenotype of Sim1 mutant mice overlaps with but is more severe than that of Brn-2 mutant mice. The Sim1 mutant hypothalamus fails to express Brn-2 during development, suggesting that a part of the phenotype of Sim1 mutant mice is due to loss of Brn-2 and that BRN-2 acts downstream of SIM1. Another bHLH-Per-ArntSim transcription factor, Arnt-2, is also involved in hypothalamic development. Arnt-2 mutant mice show a strikingly similar phenotype to the Sim1-deficient mice, consistent with the finding that SIM1 and ARNT-2 form a heterodimer complex in vivo (...

Genetic ablation of the Bsx homeodomain transcription factor in zebrafish: Impact on mature pineal gland morphology and circadian behavior

“…Several aspects of aberrant pineal function could result in the abnormal circadian rhythmicity of motor activity, obviously including disruptions in melatonin production, or alternatively photoreceptive capabilities and activity of projection neurons. Our histological analyses did not reveal any changes in brain morphology apart from that of the pineal gland, but dysfunction in other brain regions, namely the hypothalamus, 2,3,7,52–56 could play a role as well. However, the mutant phenotype reported here, that is, a zebrafish without a pineal gland, may prove to be an excellent model for studying pineal function in a context of circadian physiology in a diurnal species.…”

Circadian regulation and molecular role of the Bsx homeobox gene in the adult pineal gland

“…To determine the tissue distribution of Bsx transcripts, qRT‐PCR was performed on samples from the central nervous system and peripheral tissues obtained from adult rats sacrificed at ZT6 and ZT18 (Figure A). Bsx mRNA was detectable in the pineal gland and the hypothalamus; significantly lower levels were detected in the hypothalamus ( P < .001). Pineal Bsx mRNA showed a significant day‐night difference with higher levels at ZT18 compared with ZT6 ( P < .05), as previously indicated by RNA sequencing studies .…”

A rare case of trisomy 11q23.3-11q25 and trisomy 22q11.1-22q11.21

“…Gene knockout and overexpression experiments showed that BACE1 was related with nervous system development (Long et al, 2014). BSX, located at 11q24.1, is a DNA-binding protein and a transcriptional activator found to be essential for fetal growth and development (Chu and Ohtoshi, 2007). Alterations in AOCH, located at chromosome 11 were reported in a family with acromegaloid features, submucosal cleft palate, and diaphragmatic hernia (Stratakis et al, 2001).…”