Genomic organization and expression of the HSP70 locus in New and Old World Leishmania species C. FOLGUEIRA 1 , C. CAN ˜ AVATE 2 , C. CHICHARRO 2 and J. M. REQUENA 1 * 1 Centro de Biologı ´a Molecular «Severo Ochoa», Universidad Auto ´noma de Madrid, 28049 Madrid, Spain 2 WHO Collaborating Centre for Leishmaniasis, Servicio de Parasitologı ´a, Centro Nacional de Microbiologı ´a, Instituto de Salud Carlos III, 28220 Madrid, Spain (Received 20 June 2006; revised 31 August 2006; accepted 31 August 2006; first published online 23 October 2006) SUMMARY Heat shock is believed to be a developmental inductor of differentiation in Leishmania. Furthermore, heat shock genes are extensively studied as gene models to decipher mechanisms of gene regulation in kinetoplastids. Here, we describe the organization and expression of the HSP70 loci in representative Leishmania species (L. infantum, L. major, L. tropica, L. mexicana, L. amazonensis and L. braziliensis). With the exception of L. braziliensis, the organization of the HSP70 loci was found to be well conserved among the other Leishmania species. Two types of genes, HSP70-I and HSP70-II, were found to be present in these Leishmania species except for L. braziliensis that lacks HSP70-II gene. Polymorphisms in the HSP70 locus allow the differentiation of the Old and New World species within the subgenus Leishmania. A notable discrepancy between our data and those of the L. major genome database in relation to the gene copy number composing the L. major HSP70 locus was revealed. The temperature-dependent accumulation of the HSP70-I mRNAs is also conserved among the different Leishmania species with the exception of L. braziliensis. In spite of these differences, analysis of the HSP70 synthesis indicated that the HSP70 mRNAs are also preferentially translated during heat shock in L. braziliensis. Key words: Leishmania, HSP70, genomic organization, 3kUTR, mRNA stability, de novo synthesis of proteins. INTRODUCTION Protozoan parasites of the genus Leishmania are the aetiological agents of leishmaniasis, a group of dis- eases ranging in severity from cutaneous (CL) to mucocutaneous (MCL) and visceral (VL) forms (Murray et al. 2005). Worldwide, 2 million new cases of symptomatic disease occur each year (Desjeux, 2004). The different forms of the disease are pro- duced by different species of Leishmania. CL is mainly caused by L. tropica, L. major and L. aethio- pica in the Old World and by members of the L. mexicana (L. mexicana and L. amazonensis), L. braziliensis (L. braziliensis and L. peruviana), and L. guyanensis (L. guyanensis and L. panamensis) complexes in the New World (Anonymous, 1990). Cutaneous infection causes lesions that are normally resolved after a few months, although, depending on the causative agent, they can disseminate to mucosal surfaces. MCL is characterized by the dissemination of parasites from the skin to the naso-oropharyngeal mucosa, and is caused by infection with species of the L. braziliensis complex, especially L. braziliensis, but also by species of the L. guyanensis complex. VL is mainly caused by members of the L. donovani complex (L. donovani and L. infantum in the Old World, and L. chagasi in the New World). It is generally accepted that the clinical spectrum of leishmaniasis is governed by both parasite and host factors, but these remain poorly understood (Pearson and de Queiroz Sousa, 1996 ; Murray et al. 2005). Leishmania parasites exhibit a digenetic life-cycle. In the sandfly, they exist as extracellular promasti- gotes that are transferred to the mammalian host when the sandfly takes a bloodmeal. The parasite is phagocytosed by macrophages and, inside the acidic phagolysosomes, promastigotes differentiate to amastigotes. The mechanisms implicated in this transformation are not completely understood, although it is known that environmental factors such as pH and temperature are triggering factors of this process (Garlapati et al. 1999 ; Zilberstein and Shapira, 1994). The heat shock response is con- sidered to be of vital importance for the stage-specific differentiation of Leishmania parasites, and the genes encoding heat shock proteins (HSPs) in Leishmania have been extensively studied not only for their im- portance in the differentiation process, but also as a gene models to study gene expression in this parasite (Lee et al. 1988 ; Quijada et al. 1997; Clos and Krobitsch, 1999 ; Zilka et al. 2001). Among HSPs, HSP70 is the most highly con- served in both sequence and function; it assists a * Corresponding author : Centro de Biologı ´a Molecular ‘‘ Severo Ochoa ’’, Universidad Auto ´noma de Madrid, 28049 Madrid, Spain. Tel : +34 914978454. Fax : +34 914974799. E-mail : jmrequena@cbm.uam.es 369 Parasitology (2007), 134, 369–377. f 2006 Cambridge University Press doi:10.1017/S0031182006001570 Printed in the United Kingdom