Caracterização do gene LmHUS1 e de sua participação no fenômeno de amplificação gênica em Leishmania spp. / LmHUS1 gene characterization and its participation in the gene amplification in Leishmania spp.

AUTOR(ES)

Vinícius Santana Nunes

FONTE

IBICT - Instituto Brasileiro de Informação em Ciência e Tecnologia

DATA DE PUBLICAÇÃO

30/09/2011

RESUMO

The protozoan parasite Leishmania presents a dynamic and plastic genome in which gene amplification and chromosome translocations are common phenomena. Such plasticity hints at the necessity of dependable genome maintenance pathways. Eukaryotic cells have evolved checkpoint control systems that recognize altered DNA structures and halt cell cycle progression allowing DNA repair to take place. In these cells, the PCNA-related heterotrimeric complex formed by the proteins Hus1, Rad9, and Rad1 is known to participate in the early steps of replicative stress sensing and signaling. Here we show that the Hus1 homolog of Leishmania major is a nuclear protein that improves the cell capability to cope with replicative stress. Northern analysis and real-time PCR showed that after transfection of the gene, the selected lineage presents increase in the LmHUS1 transcripts levels and overexpression was confirmed using anti-LmHus1. Thus, overexpression of LmHus1 confers resistance to the genotoxic drugs hydroxyurea (HU) and methyl methanesulfonate (MMS) and resistance to HU correlates to reduced net DNA damage upon LmHus1 expression. Preliminary results associate the LmHus1 expression to the gene amplification phenomena in L. major. And a possible Chk1-like kinase was cloned and transfected into the parasite, the anti-Chk1 was also produced. Besides, LmHus1 mutant is presented, and the LmHUS1 gene disruption decreases its product levels, leads to serious effects on growth, and presenting an slight decrease in the resistance to genotoxic drugs. Finally, considering the hypothesis that LmHus1 participates in the damage sensing and in the control of the DNA replication threats, we hypothesized that the product of this gene acts in replication forks and is involved in DNA rearrangement s and in the amplicons generation in this parasite.

ASSUNTO(S)

1. leishmania major 2. manutenção do genoma 3. reparo de dna 4. rad9/rad1/hus1 5. amplificação gênic dna repair gene amplification genome maintenance leishmania major rad9/rad1/hus1

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