Molecular characterization of a voltage-gated potassium channel expressed in rat testis.
Molecular characterization of a voltage-gated potassium channel expressed in rat testis.
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DOI：
10.1093/molehr/6.4.303
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发表时间：
2000-04
影响因子：
4
通讯作者：
Asha Jacob;I. Hurley;Leslie O. Goodwin;George W. Cooper;S. Benoff
中科院分区：
文献类型：
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作者：
Asha Jacob;I. Hurley;Leslie O. Goodwin;George W. Cooper;S. Benoff
钾 (K(+)) 通道存在于哺乳动物睾丸和精子中。精子结合的生物素化卡里布毒素（一种 Ca(2+) 激活通道和延迟整流 K(+) 通道抑制剂）的免疫荧光检测表明，这些离子通道均匀分布在未固定的大鼠附睾精子的头部和尾部表面。基于不同类别 K(+) 通道的已知核苷酸序列，使用 PCR 引物对大鼠睾丸 RNA 进行逆转录聚合酶链反应 (RT-PCR) 分析，扩增与延迟整流 K(+) 通道同源的序列。大鼠睾丸切片的原位 RT-PCR 显示这些 K(+) 通道转录物存在于初级精母细胞和减数分裂后伸长精子细胞的细胞质中。对各种大鼠组织的 Northern 印迹分析发现了多个 K(+) 通道转录物，其中一些仅在睾丸中观察到。尝试获得全长大鼠睾丸 K(+) 通道 cDNA 序列，得到了 2693 个碱基对的组装序列，与延迟整流器 K(+) 通道 Kv1.3 具有 >90% 同源性。采用cDNA末端快速扩增方法扩增大鼠睾丸cDNA的5'端序列，但未能获得独特的序列。 DNA测序仪痕迹表明，多个相关的K(+)通道在其5'端不同，在大鼠睾丸中被扩增。
Potassium (K(+)) channels are present in both mammalian testis and spermatozoa. Immunofluorescent detection of sperm-bound biotinylated charybdotoxin, an inhibitor of Ca(2+)-activated and of delayed rectifier K(+) channels, indicated that these ion channels are uniformly distributed over the surface of both heads and tails of unfixed rat epididymal spermatozoa. Reverse transcription-polymerase chain reaction (RT-PCR) analysis on rat testis RNA with PCR primers, based on known nucleotide sequences of different classes of K(+) channels, amplified sequences homologous to delayed rectifier K(+) channels. In-situ RT-PCR on rat testis sections showed that these K(+) channel transcripts are present in the cytoplasm of primary spermatocytes and post-meiotic elongating spermatids. Northern blot analysis of various rat tissues identified multiple K(+) channel transcripts, some of which were observed only in testis. An attempt to obtain a full length rat testis K(+) channel cDNA sequence gave an assembled sequence of 2693 base pairs with >90% homology to a delayed rectifier K(+) channel, Kv1.3. A method for rapid amplification of cDNA ends was employed to amplify the 5' sequences of the rat testis cDNA but a unique sequence could not be obtained. DNA sequencer traces suggest that multiple related K(+) channels which differed at their 5' ends were amplified in rat testis.