Identification, molecular characterisation and differential expression of caveolin-1 in Trichinella spiralis maturing oocytes and embryos.
Caveolins are integral membrane proteins implicated in cholesterol homeostasis and transport, endocytosis mechanisms and regulation of signal transduction in differentiated cells. In this work a caveolin-1 gene from the nematode Trichinella spiralis (Ts-cav-1) was cloned and identified as an adult-specific antigen. For this, a cDNA library of T. spiralis 3-day-old adult worms was screened using a stage-specific cDNA-labelled probe. One positive clone contained a cDNA insert of 1427-bp and a full-length open reading frame (ORF) of 687-bp, which encodes for a 229 amino acid polypeptide with a theoretical molecular weight of 26kDa. BLAST and FASTA searches revealed a 36% and 57% identity with Caenorhabditis elegans caveolin-1, respectively. Confocal laser microscopy analysis using antibodies generated against Ts-CAV-1 protein and cross-sections of adult parasites showed that Ts-CAV-1 gradually accumulates on the surface of Trichinella oocytes and embryos, reaching a maximum at 3days p.i., and decreasing during new-born larvae (NBL) development. RT-PCR assays of parasites from 1 to 4days p.i. showed a similar gene expression profile to that observed for Ts-CAV-1 which suggests a specific developmental regulation. Free cholesterol was mainly distributed in the female germ line and it displayed increasing membrane accumulation, similar to the pattern obtained for Ts-CAV-1 protein, which suggests a temporal membrane association with Ts-CAV-1 that in turn will perform the functions mentioned above. Our results strongly indicate that Ts-cav-1 from T. spiralis plays a role in oocyte maturation and embryogenesis during development, demonstrating gender-specific expression.
Hernández-Bello
Romel
R
Department of Genetics and Molecular Biology, Centro de Investigación y Estudios Avanzados del IPN, Av I.P.N. 2508, Zacatenco 07360, México City, Mexico.
Bermúdez-Cruz
Rosa María
RM
Fonseca-Liñán
Rocío
R
García-Reyna
Patricia
P
Le Guerhier
Franck
F
Boireau
Pascal
P
Ortega-Pierres
Guadalupe
G
D000595:Amino Acid Sequence; D000818:Animals; D000947:Antigens, Helminth / Q000235:genetics; D001483:Base Sequence; D015153:Blotting, Western / Q000379:methods; D051242:Caveolin 1 / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D051244:Caveolin 3 / Q000235:genetics; D004625:Embryo, Nonmammalian / Q000378:metabolism*; D005260:Female; D015870:Gene Expression; D018507:Gene Expression Regulation, Developmental; D018613:Microscopy, Confocal; D008969:Molecular Sequence Data; D033661:Oocysts / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D017160:Trichinella spiralis / Q000196:embryology* / Q000378:metabolism*