The Entamoeba histolytica Ehcp112 gene has a distal and weak promoter.
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.
Flores-Soto
E
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.
Azuara-Liceaga
E
E
López-Camarillo
C
C
Orozco
E
E
D000818:Animals; D001483:Base Sequence; D015500:Chloramphenicol O-Acetyltransferase / Q000235:genetics; D003546:Cysteine Endopeptidases / Q000235:genetics*; D016054:DNA, Protozoan / Q000737:chemistry; D004748:Entamoeba histolytica / Q000201:enzymology / Q000235:genetics*; D005786:Gene Expression Regulation; D008969:Molecular Sequence Data; D010957:Plasmids / Q000737:chemistry; D011401:Promoter Regions, Genetic / Q000235:genetics* / Q000502:physiology; D015533:Transcriptional Activation / Q000502:physiology; D014162:Transfection
D016054:DNA, Protozoan; D015500:Chloramphenicol O-Acetyltransferase; D003546:Cysteine Endopeptidases