title: Molecular characterization of the reniform nematode C-type lectin gene family reveals a likely role in mitigating environmental stresses during plant parasitism.
abstract: The reniform nematode, Rotylenchulus reniformis, is a damaging semi-endoparasitic pathogen of more than 300 plant species. Transcriptome sequencing of R. reniformis parasitic females revealed an enrichment for sequences homologous to C-type lectins (CTLs), an evolutionarily ancient family of Ca(+2)-dependent carbohydrate-binding proteins that are involved in the innate immune response. To gain further insight as to the potential role of CTLs in facilitating plant parasitism by R. reniformis, we performed a comprehensive assessment of the CTL gene family. 5'- and 3'-RACE experiments identified a total of 11 R. reniformis CTL transcripts (Rr-ctl-1 through Rr-ctl-11) that ranged in length from 1083 to 1,194 bp and showed 93-99% identity with one another. An alignment of cDNA and genomic sequences revealed three introns with the first intron residing within the 5'-untranslated region. BLAST analyses showed the closest homologs belonging to the parasitic nematodes Heligmosomoides polygyrus and Heterodera glycines. Rr-ctl-1, -2, and -3 were expressed throughout the R. reniformis life cycle; whereas, the remaining Rr-ctl genes showed life stage-specific expression. Quantitative real time RT-PCR determined that Rr-ctl transcripts were 839-fold higher in sedentary female nematodes than the next most abundant life stage. Predicted Rr-CTL peptides ranged from 301 to 338 amino acids long, possessed an N-terminal signal peptide for secretion, and contained a conserved CLECT domain, including the mannose-binding motifs EPN and EPD and the conserved WND motif that is required for binding Ca(+2). In addition, Rr-CTL peptides harbored repeats of a novel 17-mer motif within their C-terminus that showed similarity to motifs associated with bacterial ice nucleation proteins. In situ hybridization of Rr-ctl transcripts within sedentary females showed specific accumulation within the hypodermis of the body regions exposed to the soil environment; those structures embedded within the root during parasitism did not show Rr-ctl expression. A phylogenetic analysis of the Rr-CTL CLECT domain with homologous domains from other nematode species suggested that CTLs from animal- and plant-parasitic genera may have evolved in order to play an active role in the parasitic process.
authors.affiliation: Department of Biochemistry and Molecular Biology, Mississippi State University, Mississippi State, MS 39762, USA. Electronic address: sg391@msstate.edu.
authors.forename: Satish
authors.initials: S
authors.lastname: Ganji
authors.affiliation: Department of Biochemistry and Molecular Biology, Mississippi State University, Mississippi State, MS 39762, USA; USDA-ARS, Crop Science Research Laboratory, Genetics and Precision Agriculture Research Unit, Mississippi State, MS 39762, USA. Electronic address: johnie.jenkins@ars.usda.gov.
authors.forename: Johnie N
authors.initials: JN
authors.lastname: Jenkins
authors.affiliation: Department of Biochemistry and Molecular Biology, Mississippi State University, Mississippi State, MS 39762, USA; USDA-ARS, Crop Science Research Laboratory, Genetics and Precision Agriculture Research Unit, Mississippi State, MS 39762, USA. Electronic address: martin.wubben@ars.usda.gov.
authors.forename: Martin J
authors.initials: MJ
authors.lastname: Wubben
mesh_terms: D020121:5' Untranslated Regions; D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000818:Animals; D018076:DNA, Complementary; D020224:Expressed Sequence Tags; D005260:Female; D005786:Gene Expression Regulation; D017238:Genes, Helminth; D007438:Introns; D037181:Lectins, C-Type / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D008297:Male; D008969:Molecular Sequence Data; D005810:Multigene Family; D010802:Phylogeny; D010944:Plants / Q000469:parasitology*; D017434:Protein Structure, Tertiary; D013312:Stress, Physiological / Q000235:genetics; D040521:Subcutaneous Tissue; D014431:Tylenchoidea / Q000235:genetics* / Q000254:growth & development / Q000502:physiology
chemical_list: D020121:5' Untranslated Regions; D018076:DNA, Complementary; D037181:Lectins, C-Type
keywords: C-type lectin; CTL; Ice nucleation; Parasitism; Reniform nematode; Rotylenchulus