mgr-mir-9 implicates Meloidogyne graminicola infection in rice by targeting the effector MgPDI

Abstract

MicroRNAs (miRNAs), a class of small non-coding RNAs, are crucial endogenous gene regulators in a range of animals, including plant-parasitic nematodes. Meloidogyne graminicola is an obligate sedentary endoparasite of rice and causes significant yield losses. A number of studies focused on the roles of M. graminicola effectors during the parasitic process; however, how nematode miRNAs regulate its effectors needs elucidating. In this research, we analyzed a cluster of M. graminicola miRNAs obtained at the second-stage juveniles (J2s) stage that are closely linked to the regulation of M. graminicola effectors. There are 49 767 105 total clean reads obtained from three libraries. A total of 233 known miRNAs and 21 novel miRNAs were identified. Among the known miRNAs, mgr-lin-4, mgr-mir-1, mgr-mir-100, mgr-mir-86, mgr-mir-279, mgr-mir-87, mgr-mir-71, mgr-mir-9, mgr-mir-50, mgr-mir-72, and mgr-mir-34 are the most abundant 11 miRNAs families. Moreover, the expression levels of selected miRNAs were validated by real-time quantitative PCR. We hypothesized that these miRNAs might regulate the expression of secreted effectors during the J2s stage to facilitate its infection. Consistent with this, we found that mgr-mir-9 targets MgPDI, an important M. graminicola effector mRNA. In addition to that, J2s treated with mgr-mir-9 mimics showed down-regulation of MgPDI expression and reduced reproductive ability, alluding mgr-mir-9 is involved in nematode infection. These results provide novel insight into the regulatory functions of M. graminicola miRNAs during the infection and identify miRNAs and their effector targets as potential key management targets to limit parasite survival during the early stages of infection.

Keywords

• microRNA function
• Meloidogyne graminicola
• deep sequencing
• Dual-Luciferase Reporter Assay System
• protein disulfide isomerase