Investigation on Pathophysiological Significance of Dominant Negative Isoforms of Natriuretic Peptide Receptors

c型利钠肽（CNP）利用冠酰环化酶(GC) -B作为受体，抑制血管病变增殖，促进软骨内骨化。我们已经证明，在小鼠中，单个GC-B基因产生了三个GC-B异构体（GC-B1, B2, B3），并且GC-B2缺乏一个调控区域，GC-B3由一个细胞外区域构成，仅作为GC-B1诱导的cnp诱导的细胞内cGMP水平升高的显性阴性异构体。反转录和PCR分析显示，生长板软骨中GC-B mRNA的种类以GC-B1居多，而GC-B2和GC-B3的表达水平与中枢神经系统中GC-B1 mRNA的表达水平几乎相等。这一观察结果可以解释为什么GC-B缺失小鼠最突出的表型是侏儒症，其中生长板软骨中GC-B mRNA水平作为三个亚型的总和远低于中枢神经系统。我们研究了GC-B2或GC-B3异构体是否在人体组织中表达，并证实了至少GC-B2异构体在人类心脏中表达。我们的数据表明，保留内含子8产生了一个新的GC-B异构体，内含子8包含一个框架内停止密码子，该异构体由胞外结构域、跨膜段和短细胞内段组成。预期的分子类似于人类GC-B基因GIn500的无义突变所产生的分子，这种突变会导致Maroteaux型端端粒发育不良。

C-type natriureitc peptide (CNP) utilizes guanylyl cyclase (GC) -B as a receptor to inhibit proliferative vascular lesions and to promote endochondral ossification. We have shown that three GC-B isoforms (GC-B1, B2, B3) are generated from the single GC-B gene in mice and that GC-B2 lacking a regulatory region and GC-B3 constituted by an extracellular region only serve as dominant negative isoforms against the CNP-induced increase of intracellular cGMP levels by GC-B1. A reverse transcription and PCR analysis showed that most of GC-B mRNA species in the growth plate cartilage were GC-B1, while expression levels of GC-B2 and GC-B3 are almost equal to the expression level of GC-B1 mRNA in the central nervous system. This observation could explain why the most prominent phenotype of GC-B null mice is dwarfism where the GC-B mRNA level as the sum of three isoforms in the growth plate cartilage is much less than in the cetral nervous system. We investigated if GC-B2 or GC-B3 isoforms were expressed in human tissues, and confirmed that at least GC-B2 isoform is expressed in the human heart Our data indicated that a new GC-B isoform is generated by retaining intron 8, which contains an in-frame stop codon and the isoform is consisted by an extracellular domain, a transmembrane segment, and a short intracellular segment. The expected molecule resembles the molecule generated by a nonsense mutation at GIn500 of human GC-B gene, which causes acromesomelic dysplasia, type Maroteaux.