草鱼自噬相关基因Beclin1的克隆及其在MC-LR胁迫下的表达特征

CLONING OF BECLIN1, AN AUTOPHAGY GENE, AND IT’S EXPRESSION UNDER MICROCYSTIN-LR STRESS IN GRASS CARP (CTENOPHARYNGODON IDELLA)

  • 摘要: 为评估微囊藻毒素-LR (MC-LR)对鱼类自噬的影响, 根据转录组测序结果得到的EST序列, 采用RACE技术获得了草鱼(Ctenopharygodon idella)自噬基因Beclin1 (CiBeclin1)的cDNA全长序列。该基因序列全长1590 bp, 包括1344 bp的开放阅读框, 编码447个氨基酸, 分子量为51.2 kD, 理论等电点为4.88。CiBeclin1包含1个BH3和1个ECD结构域。CiBeclin1氨基酸序列与其他物种的相似性为88%—98%, 构建的进化树显示CiBeclin1与稀有鲫(Gobiocypris rarus)的Beclin1亲缘关系最近。实时荧光定量PCR (qRT-PCR)检测结果表明, CiBeclin1在肝、肾、脾等10种不同组织中均广泛表达, 但在肝脏组织中的相对表达量最丰富, 显著高于相对表达量最低的头肾组织(P<0.05)。在不同剂量(25和100 μg MC-LR/kg BW) MC-LR胁迫24h、48h、72h和96h后, 草鱼肝脏中CiBeclin1表达量均显著低于对照组的表达水平, 研究结果表明, MC-LR能抑制草鱼CiBeclin1的表达, 但MC-LR是否在该剂量下诱导了草鱼的自噬还有待于进一步研究。

     

    Abstract: To estimate the effects of Microcystin-LR (MC-LR) on the autophagy, we cloned Beclin1 CiBeclin1 gene from grass carp (Ctenopharygodon idella) using rapid amplification of cDNA ends (RACE) based on the EST sequence. The deduced amino acid sequence of CiBeclin1 was comprised of 447 amino acids, and CiBeclin1 possessed a typical Bcl-2 homology domain 3 (BH3) and an evolutionarily conserved domain (ECD). The similarity of CiBeclin1 amino acid sequence with other species was 88%—98%, and phylogenetic analysis demonstrated that CiBeclin1 formed a clade with rare minnow Beclin1. CiBeclin1 was ubiquitously expressed in 10 tested tissues, including the liver, kidney, spleen and others, with the highest level in liver. MC-LR significantly decreased the expression of CiBeclin1 at 24, 48, 72 and 96h, indicating that CiBeclin1 gene might play a regulatory role against MC-LR toxicity. However, whether the used doses of MC-LR could induce liver autophagy needs further investigation.

     

/

返回文章
返回