Molecular cloning and chromosomal localization of the genes encoding P23k neuron-specific calcium-binding protein family.

P23k家族是一个新的EF-hand钙结合蛋白亚家族，被认为是中枢神经系统神经元中的钙传感器。我们开始在人和小鼠中寻找P23k家族的大脑成员。我们在这里报告了两个新的人类cdna和小鼠希波calcin基因。从人脑cDNA文库中获得了两个新的人类cDNA， HLP3和HLP4。两个cDNA编码一个分子量为23kDa的同源蛋白，包含4个潜在的EF-hand基序。Northern blot分析显示，这两种mrna仅在大脑中表达。HLP3mRNA分布于大部分脑区，而在大脑皮层和小脑中检测到大量的HLP3mRNA。在大脑的许多区域也发现了HLP4mRNA，在大脑皮层和小脑中检测到相对高含量的HLP4mRNA。通过对人-鼠体细胞杂交dna的不一致性分析，将HLP3和HLP4基因分别定位到2号染色体和17号染色体上。小鼠希波calcin基因从小鼠白细胞基因组文库中获得。小鼠海马蛋白基因包含3个外显子和2个内含子，长度约为8 kb。推导出的小鼠海马蛋白的氨基酸序列与大鼠和人的氨基酸序列100%相同，且其基因结构在物种间高度保守。转录终止位点位于TAG的860bp处。多个转录起始位点分布在-180bp左右。在5‘侧翼区域没有发现典型的’TATA‘盒子和’CAAT'盒子。通过Southern blot分析，阳性条带与克隆基因的阳性条带完全一致。荧光原位杂交显示小鼠海马蛋白基因定位于4号染色体D2-D3区。

P23k family is a novel EF-hand calcium-binding protein subfamily, considered to be a calcium sensor in neuron of the central nervous system. We initiated a search for the brain member of P23k family in human and mouse. We report here on two novel human cDNAs and mouse hippocalcin gene.Two novel human cDNAs, HLP3 and HLP4, were obtained from a human brain cDNA library. Both cDNA encode a homologous protein with a molecular weight of 23kDa, containing four potential EF-hand motifs. Northern blot analysis revealed that both mRNAs were expressed only in the brain. The HLP3 mRNA was distributed to most of the brain regions, high amounts of the HLP3mRNA were detected in the cerebral cortex and cerebellum. The HLP4mRNA was also found in many regions of the brain, relatively high amounts of the HLP4mRNA were detected in the cerebral cortex and cerebellum. By discordancy analysis using DNAs from human-rodent somatic cell hybrids, the HLP3 and HLP4 genes were mapped to chromosome 2 and 17, respectively.The mouse hippocalcin gene was obtained from a mouse leukocyte genomic library. The mouse hippocalcin gene contains three exons and two introns, which spans approximately 8 kb. Deduced amino acid sequence of mouse hippocalcin is 100% identical to those of the rat and human and structures of hippocalcin genes are highly conserved across species. The transcription termination site was assigned at 860bp from TAG.The multiple transcription initiation sites were assigned at the positions around-180bp. Neither a canonical 'TATA' box, and 'CAAT' box was found in the 5'-flanking region. By Southern blot analysis, the positive bands exactlycoincided with those expected from the cloned gene. Fluorescence in situ hybridization revealed the mouse hippocalcin gene mapped to chromosome 4, region D2-D3.