title: Characterization of pneumocystis major surface glycoprotein gene (msg) promoter activity in Saccharomyces cerevisiae.
abstract: Major surface glycoprotein (Msg), the most abundant cell surface protein of Pneumocystis, plays an important role in the interaction of this opportunistic pathogen with host cells, and its potential for antigenic variation may facilitate evasion of host immune responses. In the present study, we have identified and characterized the promoter region of msg in 3 species of Pneumocystis: P. carinii, P. jirovecii, and P. murina. Because Pneumocystis cannot be cultured, promoter activity was measured in Saccharomyces cerevisiae, a related fungus, using a yeast vector modified to utilize the gene coding for Renilla luciferase as a reporter gene. The 5'-flanking sequences of msg from all three Pneumocystis species showed considerable promoter activity, with increases in luciferase activity up to 15- to 44-fold above baseline. Progressive deletions helped define an ∼13-bp sequence in each Pneumocystis species that appears to be critical for promoter activity. Electrophoretic mobility shift analysis using P. carinii-specific msg promoter sequences demonstrated binding of nuclear proteins of S. cerevisiae. The 144-bp 5'-flanking region of P. murina msg showed 72% identity to that of P. carinii. The 5'-flanking region of P. jirovecii msg showed 58 and 61% identity to those of P. murina and P. carinii, respectively. The msg promoter is a good candidate for inclusion in a construct designed for genetic manipulation of Pneumocystis species.
authors.affiliation: Critical Care Medicine Department, NIH Clinical Center, NIH, Bethesda, Maryland, USA.
authors.forename: Geetha
authors.initials: G
authors.lastname: Kutty
authors.forename: Robert
authors.initials: R
authors.lastname: Shroff
authors.forename: Joseph A
authors.initials: JA
authors.lastname: Kovacs
mesh_terms: D001483:Base Sequence; D005656:Fungal Proteins / Q000235:genetics* / Q000378:metabolism; D006023:Glycoproteins / Q000235:genetics* / Q000378:metabolism; D008565:Membrane Proteins / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D011010:Pneumocystis / Q000235:genetics* / Q000378:metabolism; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D012441:Saccharomyces cerevisiae / Q000378:metabolism; D014158:Transcription, Genetic
chemical_list: D005656:Fungal Proteins; D006023:Glycoproteins; D008565:Membrane Proteins
references.citation: Emerg Infect Dis. 2002 Sep;8(9):891-6
references.pmid: 12194762
references.citation: DNA Res. 1995;2(2):77-88
references.pmid: 7584051
references.citation: J Infect Dis. 1999 Dec;180(6):1969-78
references.pmid: 10558954
references.citation: Mol Microbiol. 2001 Oct;42(1):183-93
references.pmid: 11679077
references.citation: Fungal Genet Biol. 2007 Sep;44(9):905-19
references.pmid: 17320432
references.citation: EMBO J. 1996 May 1;15(9):2227-35
references.pmid: 8641288
references.citation: J Eukaryot Microbiol. 1994 Sep-Oct;41(5):84S
references.pmid: 7804279
references.citation: J Infect Dis. 2001 Nov 15;184(10):1358-62
references.pmid: 11679931
references.citation: J Biol Chem. 1993 Mar 15;268(8):6034-40
references.pmid: 8449961
references.citation: Mol Microbiol. 1997 Jul;25(1):147-60
references.pmid: 11902717
references.citation: BMC Genomics. 2009 Aug 07;10:367
references.pmid: 19664205
references.citation: J Clin Invest. 1991 Jan;87(1):163-70
references.pmid: 1985093
references.citation: DNA Res. 1998 Apr 30;5(2):77-85
references.pmid: 9679195
references.citation: J Immunol. 1988 Mar 15;140(6):2023-31
references.pmid: 3257995
references.citation: Infect Immun. 1994 Aug;62(8):3092-101
references.pmid: 7518806
references.citation: Infect Immun. 1989 Jul;57(7):2149-57
references.pmid: 2786505
references.citation: Microbiology (Reading). 2004 May;150(Pt 5):1153-1165
references.pmid: 15133075
references.citation: J Infect Dis. 1992 Nov;166(5):1113-23
references.pmid: 1402023
references.citation: Mycologia. 2004 May-Jun;96(3):429-38
references.pmid: 21148866
references.citation: J Infect Dis. 1995 Jun;171(6):1563-8
references.pmid: 7769293
references.citation: J Eukaryot Microbiol. 1997 Nov-Dec;44(6):10S-11S
references.pmid: 9508404
references.citation: Mol Microbiol. 2008 May;68(3):547-59
references.pmid: 18394144
references.citation: Mol Biochem Parasitol. 1991 Mar;45(1):183-4
references.pmid: 1711155
references.citation: DNA Cell Biol. 1996 Nov;15(11):989-99
references.pmid: 8945640
references.citation: Nat Methods. 2012 Jul;9(7):671-5
references.pmid: 22930834
references.citation: JAMA. 2009 Jun 24;301(24):2578-85
references.pmid: 19549975
references.citation: N Engl J Med. 2004 Jun 10;350(24):2487-98
references.pmid: 15190141
references.citation: J Eukaryot Microbiol. 1993 Nov-Dec;40(6):733-41
references.pmid: 8292993