Small RNA and Transcriptome Analysis of Cotton (Gossypium hirsutum L.) Susceptibility, Resistance, and Hypersensitivity to Reniform Nematodes (Rotylenchulus reniformis)

Abstract

Reniform nematodes (RN) are semi-endoparasitic nematode species causing significant yield loss in cotton, particularly in the eastern US cotton belt. Successful RN parasitism is contingent on establishment of a syncytium, which serves as the sole nutrient source on which RN live. The syncytia are hypertrophied, multinucleate root cells with enlarged nuclei and dense cytoplasm, which resulted from the breakdown of cell walls between initial feeding cells and neighboring cells. It is believed that nematode secretions injected through their stylet, a specialized needle-like structure mouthpart, are essential in syncytium initiation and maintenance. In response to nematode infestation, plant resistance relies on the coordination of different resistance mechanisms including specific resistance genes or proteins, several plant hormone pathways, and reactive oxygen species (ROS) that are generated in response to nematode attack. These resistance-related elements crosstalk to each other and can be seen as an integrated signaling network regulated by transcription factors and small RNAs (sRNAs) at the transcriptional (epigenetic), posttranscriptional, and/or translational levels. However, little is known about the mechanisms involved in host responses to RN infestation. The overall objective of this project is to identify and characterize such regulatory networks in cotton root responses to RN infestation with the following specific objectives: 1) To determine transcriptom and sRNA expression in cotton roots with different levels of RN resistance with and without RN infestation, 2) to correlate the identified sRNAs and transcriptome expression profiles and build potential regulatory networks that are important in mediating different levels of resistance to RN in cotton roots. To accomplish these objectives, both transcriptome and sRNA libraries were constructed from RN susceptible, resistant, and hypersensitive genotypes of cotton seedlings, with and without RN infestation. The expression of transcriptome was analyzed to detect RN responsive genes and important gene differences between genotypes with varied levels of RN resistance. A number of known genes involved in generic plant-nematode interactions, as well as genes that are newly identified to be involved in cotton-RN interactions, were detected in this study. For the second objective, conserved miRNAs and lineage-specific miRNAs were identified, and the target sequences of the identified differentially expressed miRNAs were determined from the custom assembled transcriptome data. SRNA regulatory networks involving miRNAs and their negatively regulated target genes were suggested to play important roles in RN pathogenesis in cotton roots. Taken together, the work in this study identified genes and sRNAs that are important in plant responses to RN infestation and in the genotypic variations for RN resistance, and these results will set a foundation for future research towards understanding the resistance mechanisms to RN in cotton.