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Identification of Novel and Conserved MicroRNAs of Root-knot Nematode Using a Deep-sequencing Approach

ABSTRACT: 111,556,278 high-quality reads were obtained by a deep-sequencing approach that show an exact match to the genome of Meloidogyne incognita from the library of J2 juveniles of M. incognita (Mi). Based on these Solexa reads, 89 M. incognita microRNA genes were identified, which grouped into 67 non-redundant miRNAs with mature sequences. All of these candidate miRNAs were confirmed by qRT-PCR, and 26 of them could be detected in the library of the galls of cucumber root infected by M. incognita (GC). MiR-100 was found in a cluster with let-7, which is similar with B. malayi, a human parasitic nematode. Based on the results of deep sequencing, the expression of miR-100 was much more abundant than that of let-7, which indicated that they may not be co-expressed. The ortholog of let-7, a key regulator that controls the nematode from L4 to adult in C. elegans, could be frequently sequenced in the GC library, the later stages of development of M. incognita, while it had a relative low expression level in J2, which indicated that let-7 may have a similar role in the development regulation in plant parasitic nematodes. Frequently sequenced microRNAs, including miR-71, miR-100 and miR-124, should play an important role in the growth and proliferation of M. incognita.

ORGANISM(S): Capsicum annuum Meloidogyne incognita

PROVIDER: GSE24833 | GEO | 2011/07/21

SECONDARY ACCESSION(S): PRJNA132045

REPOSITORIES: GEO

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Project description:The root knot nematode, Meloidogyne incognita, is an obligate parasite that causes significant damage to a broad range of host plants. Infection is associated with secretion of proteins surrounded by proliferating cells. Many parasites are known to secrete effectors that interfere with plant innate immunity, enabling infection to occur; they can also release pathogen-associated molecular patterns (PAMPs, e.g., flagellin) that trigger basal immunity through the nematode stylet into the plant cell. This leads to suppression of innate immunity and reprogramming of plant cells to form a feeding structure containing multinucleate giant cells. Effectors have generally been discovered using genetics or bioinformatics, but M. incognita is non-sexual and its genome sequence has not yet been reported. To partially overcome these limitations, we have used mass spectrometry to directly identify 486 proteins secreted by M. incognita. These proteins contain at least segmental sequence identity to those found in our 3 reference databases (published nematode proteins; unpublished M. incognita ESTs; published plant proteins). Several secreted proteins are homologous to plant proteins, which they may mimic, and they contain domains that suggest known effector functions (e.g., regulating the plant cell cycle or growth). Others have regulatory domains that could reprogram cells. Using in situ hybridization we observed that most secreted proteins were produced by the subventral glands, but we found that phasmids also secreted proteins. We annotated the functions of the secreted proteins and classified them according to roles they may play in the development of root knot disease. Our results show that parasite secretomes can be partially characterized without cognate genomic DNA sequence. We observed that the M. incognita secretome overlaps the reported secretome of mammalian parasitic nematodes (e.g., Brugia malayi), suggesting a common parasitic behavior and a possible conservation of function between metazoan parasites of plants and animals.

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Identification of Novel and Conserved MicroRNAs of Root-knot Nematode Using a Deep-sequencing Approach

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