Pneumocystis carinii expresses an active Rtt109 histone acetyltransferase.
Species in the genus Pneumocystis can cause severe pneumonia in immune-compromised hosts. The identification of specific targets present in Pneumocystis species, but lacking in mammalian hosts, is paramount to developing new means to treat this infection. One such potential protein is Rtt109, which is a type of histone acetyltransferase (HAT) required for DNA replication in fungi, but not found in mammals. Sequence orthologues of Rtt109 are present in other fungi, but are absent in mammals, making it a potential pan-specific target against medically relevant fungi. Accordingly, we sought to identify the presence of an Rtt109 in P. carinii. A Pneumocystis carinii (Pc) Rtt109 165-bp partial sequence was initially identified from the incomplete P. carinii genome database. Subsequently, a full-length, 1,128-bp cDNA with homology to Saccharomyces cerevisiae Rtt109 (39% Basic Local Alignment Search Tool (BLASTP)) was cloned and characterized. Sequence analysis of PcRtt109 indicated that the P. carinii molecule contains the putative catalytic aspartate present in yeast. We further demonstrated that the PcRtt109 expressed in rtt109Δ S. cerevisiae cells restored H3-K56 acetylation and the sensitivity toward DNA-damaging agents of rtt109Δ mutant cells. Purified PcRtt109 had the ability to acetylate lysine-56 of histone H3, similar to the ability of Schizosaccharomyces pombe Rtt109 protein. The site-directed mutagenesis of PcRtt109 D84A, a potential regulatory site in the Rtt109 HAT family, abolished H3 acetylation, whereas a DD218/219AA mutation that compromised the activity of ScRtt109 had little effect, demonstrating similarities and differences in Pneumocystis PcRtt109 compared with yeast Saccharomyces cerevisiae Rtt109. These results indicate that P. carinii contains an Rtt109 HAT molecule, and represent the complete identification and characterization of a HAT molecule from this important opportunistic fungal pathogen.
Kottom
Theodore J
TJ
Thoracic Diseases Research Unit, Department of Medicine and Department of Biochemistry, Mayo Clinic, Rochester, MN 55905, USA.
Han
Junhong
J
Zhang
Zhiguo
Z
Limper
Andrew H
AH
D000595:Amino Acid Sequence; D000818:Animals; D015966:Gene Expression Regulation, Fungal; D051548:Histone Acetyltransferases / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism; D006657:Histones / Q000737:chemistry; D007107:Immune System; D008168:Lung / Q000382:microbiology; D008969:Molecular Sequence Data; D045363:Pneumocystis carinii / Q000378:metabolism*; D051381:Rats; D011994:Recombinant Proteins / Q000737:chemistry; D012441:Saccharomyces cerevisiae / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000096:biosynthesis* / Q000235:genetics; D017386:Sequence Homology, Amino Acid
D006657:Histones; D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins; D051548:Histone Acetyltransferases; C517120:Rtt109 protein, S cerevisiae