Evaluation of the amino-acid polymorphism identified in the insulin gene transcription factor A1

两个基本的螺旋-环-螺旋（bHLH）转录因子E47和E12作为二聚体复合物的一部分参与胰岛素基因的表达，它们与顺式作用基序E-box相互作用。虽然两者都是通过选择性剪接从单一基因（E2 A）产生的，但两者之间这些bHLH区域的结构差异表明两种bHLH蛋白可能在某些功能上不同。我们最近发现了亮氨酸拉链（LZ）基序内高度保守区域的氨基酸（aa）多态性。有趣的是，氨基酸缺失的位置和模式与先前在人类对应物E47（E12）中描述的氨基酸多态性相同[Cell 60（1990）547-555]，即谷氨酰胺的插入或缺失。（C57BL/6 ob/ob、C57BL/6J、C57BL/KsJ、ICR、BALB/c ByJ、DBA/2J和C3H/HeJ）。的LZ区域 ...更多信息 克隆AI cDNA（nt 31 -401）并进行DNA测序。在我们检测的7个菌株中，6个具有谷氨酰胺（-）-A1型，这表明这种A1型在菌株数量方面相当普遍。只有C3 H菌株具有谷氨酰胺（+）-型A1。由于已知C3 H菌株是相对高胰岛素血症的，因此提出了可能的生理意义。然而，这两种类型的A1，当在CHO细胞中表达时，显示出相同的DNA结合活性，这表明它们至少在CHO细胞中保留了相同的DNA结合活性。为了阐明E47和E12的个体意义，我们还研究了它们的同源物（分别为A1和KA 1）在小鼠组织和细胞系中的mRNA表达比率。使用逆转录聚合酶链反应和特定的寡脱氧核糖核苷酸杂交技术的测量结果显示，A1和kA 1 mRNA在所有检查的组织中普遍表达。然而，它们的比例不同：例如骨骼肌，2.2 - <plus-minus>0.3（平均<plus-minus>SE），脾脏，2.0 - <plus-minus>0.2，胰岛细胞，1.2 - <plus-minus>0.2。研究的细胞系中的A1 kA 1比率与其原始组织中的A1 kA 1比率相似。在小鼠中观察到的E47和E12同源物mRNA表达的普遍性为它们参与广泛的转录调控提供了支持。另一方面，A1/KA 1表达比例的变化支持A1（E47）和KA 1（E12）在这些E2 A基因编码的bHLH p Less的功能中各自具有某些独特作用的观点。

Two basic helix-loop-helix (bHLH) transcription factors, E47 and E12, are involved in the insulin geneexpression as part of dimeric complexes which interact with the cis-acting motif, E-box. Although both generated from a single gene (E2A) by means of alternative splicing, the structural difference in these bHLH regions between the two suggests that two bHLH proteins may differ in some of their functions. We reccuntly indentified an amino acid (aa) polymorphism in the highly conserved region within the leucine zipper (LZ) motif. Interestingly, the location and pattern of aa delections are identical to those previously described for the aa polymorphism within the human counterpart, E47 (E12) [Cell60 (1990) 547-555], which was insertion or deletion of glutamine.To explore the possible physiological meaning of the polymorphism, we surveyed seven mouse strains (C57BL/6 ob/ob, C57BL/6J,C57BL/KsJ,ICR,BALB/c ByJ,DBA/2J and C3H/HeJ) in terms of the type of this polymorphism. The LZ regions of … More the AI cDNAs (nt31-401) were cloned and DNA-sequenced. Among the seven strains that we examined, six had the glutamine (-)-type A1, suggesting that this type of A1 is rather prevalent in terms of numbers of strains. Only the C3H strain had the glutamine (+)-type A1. Because the C3H strain is known to be relatively hyperinsulinemic, the possible physiological significance was suggested. However, the two types of A1, when expressed in CHO cells, revealed the same DNA-binding activity, suggesting that they retains the same DNA-binding activity at least in CHO cells.To elucidate the individual implications of E47 and E12, we also investigated the mRNA expression ratios of their homologues (A1 and KA1, respectively) in mouse tissues and cell lines. Measurement using reverse-transcription polymerase chain reaction and a specific oligodeoxyribonucleotide hybridization technique revealed that both the A1 and kA1 mRNAs were ubiquitously expressed in all tissues examined. However, their ratios varied : e.g.skeletal muscle, 2.2<plus-minus>0.3 (mean<plus-minus>SE), spleen, 2.0<plus-minus>0.2, pancreatic islet cells, 1.2<plus-minus>0.2. The A1 kA1 ratios in the cell lines investigated were similar to those of their original tissues. The ubiquity in mRNA expression observed for both the E47 and E12 homologues in mouse provides support for their involvement in a broad range of transcriptional regulation. The variation in the A1/kA1 expression ratios, on the other hand, supports the idea that A1 (E47) and KA1 (E12) each have some unique roles in the functions of these E2A geneencoded bHLH p Less