The distribution pattern of proliferating cell nuclear antigen in the nuclei of Leishmania donovani

• Published in: Microbiology (Society for General Microbiology) 2009-11, Vol.155 (11), p.3748-3757
• Main Authors: Kumar, Devanand, Minocha, Neha, Rajanala, Kalpana, Saha, Swati
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Biochemistry. Physiology. Immunology. Molecular biology; Biological and medical sciences; Cell Cycle; Cell Nucleus - metabolism; Cloning, Molecular; DNA Replication; DNA, Protozoan - biosynthesis; Fundamental and applied biological sciences. Psychology; Leishmania donovani; Leishmania donovani - cytology; Leishmania donovani - genetics; Leishmania donovani - metabolism; Molecular Sequence Data; Proliferating Cell Nuclear Antigen - genetics; Proliferating Cell Nuclear Antigen - metabolism; Protozoa; Sequence Alignment
• ISSN: 1350-0872; 1465-2080
• DOI: 10.1099/mic.0.033217-0

1 Department of Microbiology, University of Delhi South Campus, Benito Juarez Road, New Delhi 110021, India 2 National Institute of Immunology, Aruna Asaf Ali Marg, New Delhi 110067, India DNA replication in eukaryotes is a highly conserved process marked by the licensing of multiple origins, with pre-replication complex assembly in G1 phase, followed by the onset of replication at these origins in S phase. The two strands replicate by different mechanisms, and DNA synthesis is brought about by the activity of the replicative DNA polymerases Pol and Pol . Proliferating cell nuclear antigen (PCNA) augments the processivity of these polymerases by serving as a DNA sliding clamp protein. This study reports the cloning of PCNA from the protozoan Leishmania donovani , which is the causative agent of the systemic disease visceral leishmaniasis. PCNA was demonstrated to be robustly expressed in actively proliferating L. donovani promastigotes. We found that the protein was present primarily in the nucleus throughout the cell cycle, and it was found in both proliferating procyclic and metacyclic promastigotes. However, levels of expression of PCNA varied through cell cycle progression, with maximum expression evident in G1 and S phases. The subnuclear pattern of expression of PCNA differed in different stages of the cell cycle; it formed distinct subnuclear foci in S phase, while it was distributed in a more diffuse pattern in G2/M phase and post-mitotic phase cells. These subnuclear foci are the sites of active DNA replication, suggesting that replication factories exist in Leishmania , as they do in higher eukaryotes, thus opening avenues for investigating other Leishmania proteins that are involved in DNA replication as part of these replication factories. Correspondence Swati Saha ss5gp{at}yahoo.co.in Abbreviations: DAPI, 4',6-diamidino-2-phenylindole; ORC, origin recognition complex; PCNA, proliferating cell nuclear antigen; pre-RC, pre-replication complex The GenBank/EMBL/DDBJ accession number for the LdPCNA sequence reported in this paper is GQ249893. A CLUSTAL W analysis of LdPCNA and PCNAs from various trypanosomatids is available with the online version of this paper.