RESUMEN
SUMMARYNewborn larvae (NBL) and adult (Ad) stage-specifically expressed genes or members of gene families of Trichinella spiralis were identified by suppression subtractive hybridization (SSH). Six cDNA clones were identified as NBL stage-specific, including 1 member of the T. spiralis gene family encoding glutamic acid-rich proteins, 2 clones encoding novel serine proteases, 2 closely related clones encoding proteins that are members of a deoxyribonuclease II (DNase II)-like family and 1 clone with no similarity to known genes. Four stage-specific clones encoding homologues of retinoid X receptor, caveolin, C2H2 type zinc finger protein and a putative protein with no homology to known sequences were obtained from 3-day-old adult worms. One gene specifically up-regulated in the 5-day-old adult worms encoding a putative cuticle collagen was also identified.
Asunto(s)
Regulación del Desarrollo de la Expresión Génica , Estadios del Ciclo de Vida/genética , Trichinella spiralis/crecimiento & desarrollo , Trichinella spiralis/genética , Animales , ADN Complementario/metabolismo , Perfilación de la Expresión Génica/veterinaria , Larva/genética , Larva/fisiología , Ratones , Datos de Secuencia Molecular , Hibridación de Ácido Nucleico/métodos , Ratas , Trichinella spiralis/metabolismoRESUMEN
A cDNA library from Trichinella spiralis adults 3 days post-infection was screened with a cDNA probe, designated T 54, derived from a newborn larvae subtracted cDNA library. Sequence analysis showed that the positive clone contained a cDNA insert of 1464 bp in length with a single open reading frame of 1290 bp, which encoded a protein of 429 amino acids with a putative molecular mass of 49.9 k Da. Database analysis predicted the deduced protein had a leucine zipper motif and an FYVE zinc finger domain. The recombinant fusion protein was expressed and rabbit anti-recombinant protein sera reacted with a single peptide migrating at approximately 55 k Da in crude worm extract from muscle larvae, adults and newborn larvae stages.
Asunto(s)
Trichinella spiralis/genética , Dedos de Zinc/genética , Secuencia de Aminoácidos , Animales , Western Blotting , Clonación Molecular , ADN Complementario/química , ADN Complementario/genética , Biblioteca de Genes , Leucina Zippers/genética , Ratones , Datos de Secuencia Molecular , Hibridación de Ácido Nucleico , Ratas , Proteínas Recombinantes/genética , Trichinella spiralis/aislamiento & purificación , Triquinelosis/parasitologíaRESUMEN
The aim of the study was to isolate genes coding for stage-specific antigens of T. spiralis. Such antigens may then be associated with local and systemic immune responses against adult T. spiralis. Recombinant clones were obtained with an adult stage specific probe from a cDNA library of three-day old adult T. spiralis. Several cDNA clones encoding the same peptide were identified and their stage specificity was confirmed by northern blot analysis. Three independent clones were fully sequenced, and the resulting sequence found to code for a 487 amino acid peptide with a deduced molecular weight of approximately 55 kDa. Sequence analysis showed that the 55 kDa peptide contained putative DNA binding motifs, suggesting that this protein may be involved in transcriptional regulation during the early development of the parasite.