Dirofilaria immitis: ultrastructural localization, molecular characterization, and analysis of the expression of p27, a small heat shock protein homolog of nematodes.

A cDNA fragment encoding the complete coding region of a 27-kDa protein (p27) of Dirofilaria immitis was cloned. Antibody to the recombinant p27 bound to hypodermal tissues of third (L3) and fourth stage larvae (L4) of D. immitis and to both the hypodermis and the cuticle of L3s of Onchocerca volvulus, as visualized by immunoelectronmicroscopy. The deduced amino acid sequence of the central and C-terminal regions of p27 (amino acids S83 to H222) is 18-36% identical to members of the sHsp/alpha-crystallin family of proteins. The homologous region is thought to be responsible for the molecular chaperone activity of members of this family. The p27 cDNA does not encode a hydrophobic signal peptide. At least two homologous yet distinct p27 genes were identified in the D. immitis genome by Southern hybridization using the p27 cDNA as a probe. The p27 transcript was 0.9 kb in length on Northern blots. The expression of p27 in L3s of D. immitis was neither upregulated by heat shock (43 degrees C) nor by incubation at the physiologic temperature of 37 degrees C. Pulse-labeling experiments of both D. immitis and Brugia malayi L3s during the L3-L4 molt in vitro showed that synthesis of p27 is also not upregulated during this developmental phase. However, p27 is expressed constitutively throughout the D. immitis L3-L4 molt and therefore by both larval stages. In addition, both female and male adult worms of this species express p27 constitutively. P27, or an allomorph thereof, was detected in each of nine species representing four nematode superfamilies, thus indicating that this molecule is ubiquitous within the phylum Nematoda. In view of the hypodermal localization of p27, its constitutive expression, and its retention among nematodes, the function of this protein in essential housekeeping roles such as that of molecular chaperone during the molting process is discussed.