Gonzalez-Cadavid N F, Burnett A L, Magee T R, Zeller C B, Vernet D, Smith N, Gitter J, Rajfer J
Department of Urology, UCLA School of Medicine, Harbor-UCLA Medical Center, Torrance, California 90509, USA.
Biol Reprod. 2000 Sep;63(3):704-14. doi: 10.1095/biolreprod63.3.704.
Penile erection is mediated by nitric oxide (NO) synthesized by the neuronal nitric oxide synthase (nNOS). In the rat penis, the main nNOS mRNA variant, PnNOS, differs from cerebellar nNOS (CnNOS) by a 102 base pair insert encoding a 34-amino acid sequence. In the mouse, two nNOS mRNAs have been identified: nNOSalpha, encoding a 155-kDa protein, and an exon 2-deletion variant, nNOSbeta, encoding a 135-kDa protein that lacks a domain where a protein inhibitor of nNOS (PIN) binds. We wished to determine whether PnNOSalpha and beta are expressed in the rat penis and are located in the nerves and whether the beta form persists in the potent nNOS knock-out mouse (nNOS( big up tri, open big up tri, open)). A PnNOS antibody against the insert common to both PnNOSalpha and beta detected the expected 155-kDa protein in PnNOSalpha-transfected cells. This antibody, and the one common to PnNOS/CnNOS, showed (on Western blots) the 155- and 135-kDa nNOS variants in rat penile tissue during development and aging. PnNOSalpha mRNA and its subvariants were found as the main nNOS in the penile corpora, the cavernosal nerve, and the pelvic ganglia, with lower levels of PnNOSbeta mRNA. In tissue sections, PnNOS protein was immunodetected in the penile nerve endings in the rat and in the nNOS wild-type and nNOS( big up tri, open big up tri, open) mice. An antibody against the sequence encoded by exon 2 did not react (on Western blots) with the 135-kDa band, which confirms that this protein is the beta form. In conclusion, both PnNOSalpha and beta are expressed in the rat penis at all ages and are located in the nerves. The beta form may allow nitric oxide synthesis during erection to be partially insensitive to PIN. The residual expression of PnNOS, and possibly CnNOS, in the penis of the nNOS( big up tri, open big up tri, open) mouse occurs through transcription of the beta mRNA, and this may explain the retention of erectile function when the expression of nNOSalpha is disrupted.
阴茎勃起由神经元型一氧化氮合酶（nNOS）合成的一氧化氮（NO）介导。在大鼠阴茎中，主要的nNOS mRNA变体PnNOS与小脑nNOS（CnNOS）的区别在于有一个102个碱基对的插入片段，该片段编码一个34个氨基酸的序列。在小鼠中，已鉴定出两种nNOS mRNA：编码155 kDa蛋白的nNOSα和编码135 kDa蛋白的外显子2缺失变体nNOSβ，后者缺少nNOS蛋白抑制剂（PIN）结合的结构域。我们希望确定PnNOSα和β是否在大鼠阴茎中表达并位于神经中，以及β形式在有效的nNOS基因敲除小鼠（nNOS(大上三角，开放大上三角，开放)）中是否持续存在。一种针对PnNOSα和β共有的插入片段的PnNOS抗体在转染了PnNOSα的细胞中检测到预期的155 kDa蛋白。该抗体以及PnNOS/CnNOS共有的抗体在蛋白质印迹上显示了大鼠阴茎组织在发育和衰老过程中的155 kDa和135 kDa的nNOS变体。PnNOSα mRNA及其亚变体在阴茎海绵体、海绵体神经和盆腔神经节中作为主要的nNOS被发现，而PnNOSβ mRNA水平较低。在组织切片中，在大鼠以及nNOS野生型和nNOS(大上三角，开放大上三角，开放)小鼠的阴茎神经末梢中免疫检测到了PnNOS蛋白。一种针对外显子2编码序列的抗体在蛋白质印迹上不与135 kDa条带反应，这证实该蛋白是β形式。总之，PnNOSα和β在大鼠阴茎的所有年龄段均有表达且位于神经中。β形式可能使勃起过程中的一氧化氮合成对PIN部分不敏感。在nNOS(大上三角，开放大上三角，开放)小鼠的阴茎中，PnNOS以及可能的CnNOS的残留表达是通过β mRNA的转录发生的，这可能解释了nNOSα表达被破坏时勃起功能的保留。
