Flores-Soto E, Azuara-Liceaga E, López-Camarillo C, Orozco E
Posgrado en Ciencias Genómicas de la Universidad Autónoma de la Ciudad de México, Mexico D.F. Ave. San Lorenzo # 290, Col. del Valle, Mexico, D.F. 03100, Mexico.
Exp Parasitol. 2005 Jul;110(3):286-91. doi: 10.1016/j.exppara.2005.03.003. Epub 2005 Apr 9.
Ehcp112 encodes the Entamoeba histolytica EhCP112 cysteine protease that is part of the EhCPADH complex. By in silico analysis we identified putative transcription factor-binding sites along 837 bp upstream the Ehcp112 gene ATG codon. A TATA-like motif (TATATAAA) was located at -36 to -29 bp, a GAAC box (GAACC) was found at -10 to -14 bp and an Inr sequence (TTCAAC) at -8 to -2 bp. These tripartite promoter elements are in non-canonical positions, downstream the transcription initiation site (-280 bp). We cloned four Ehcp112 promoter fragments in pBSCAT-ACT plasmid to obtain pI (355 bp), pII (681 bp), pIII (833 bp), and pIV (554 bp) constructs. In transfected trophozoites, only pIII drove CAT activity with 44% efficiency in relation to actin promoter activity. Our results showed the presence of a distal and weak promoter in the Ehcp112 gene. The active DNA region is inside the open reading frame of the Ehrab B gene, suggesting that expression of both genes could be coordinately regulated.
Ehcp112编码溶组织内阿米巴的EhCP112半胱氨酸蛋白酶，它是EhCPADH复合物的一部分。通过计算机分析，我们在Ehcp112基因ATG密码子上游837 bp处鉴定出假定的转录因子结合位点。一个类似TATA的基序（TATATAAA）位于-36至-29 bp处，一个GAAC框（GAACC）在-10至-14 bp处被发现，一个Inr序列（TTCAAC）在-8至-2 bp处。这些三方启动子元件处于非典型位置，在转录起始位点（-280 bp）下游。我们将四个Ehcp112启动子片段克隆到pBSCAT-ACT质粒中，以获得pI（355 bp）、pII（681 bp）、pIII（833 bp）和pIV（554 bp）构建体。在转染的滋养体中，只有pIII驱动CAT活性，相对于肌动蛋白启动子活性，效率为44%。我们的结果表明Ehcp112基因中存在一个远端且较弱的启动子。活性DNA区域位于Ehrab B基因的开放阅读框内，表明这两个基因的表达可能受到协同调控。
