Silencing the Myosin Regulatory Light Chain Gene sqh Reduces Cold Hardiness in Ophraella communa LeSage (Coleoptera: Chrysomelidae)

SIMPLE SUMMARY: Cold hardiness is critical to the ability of insects to survive in cold climates and expand their geographical distribution. The molecular mechanisms underlying insect cold tolerance have been well-studied, but many potential genes that may impact these responses, including MRLC-sqh, have not been thoroughly evaluated. We first cloned and characterized the MRLC-sqh from Ophraella communa, an effective biological control agent of Ambrosia artemisiifolia, and found that the protein sequence was highly conserved across various Coleoptera insects. The relative expression of MRLC-sqh was tissue- and stage-specific, with high levels of expression in the gut and pupal stage of O. communa. In addition, the expression of MRLC-sqh was shown to decrease after cold shock between 10 and 0 °C and ascend between 0 and −10 °C, but these did not show a positive association between MRLC-sqh expression and cold stress. Silencing of MRLC-sqh prolonged the chill-coma recovery time in these beetles, suggesting their cold hardiness was reduced in the absence of this protein. Therefore, these results indicate that MRLC-sqh may be partly responsible for the regulation of cold-tolerance responses in insects. ABSTRACT: Ambrosia artemisiifolia is a noxious invasive alien weed, that is harmful to the environment and human health. Ophraella communa is a biocontrol agent for A. artemisiifolia, that was accidentally introduced to the Chinese mainland and has now spread throughout southern China. Recently, we found that upon artificial introduction, O. communa can survive in northern China as well. Therefore, it is necessary to study the cold hardiness of O. communa. Many genes have been identified to play a role in cold-tolerance regulation in insects, but the function of the gene encoding non-muscle myosin regulatory light chain (MRLC-sqh) remains unknown. To evaluate the role played by MRLC-sqh in the cold-tolerance response, we cloned and characterized MRLC-sqh from O. communa. Quantitative real-time PCR revealed that MRLC-sqh was expressed at high levels in the gut and pupae of O. communa. The expression of MRLC-sqh was shown to decrease after cold shock between 10 and 0 °C and ascend between 0 and −10 °C, but these did not show a positive association between MRLC-sqh expression and cold stress. Silencing of MRLC-sqh using dsMRLC-sqh increased the chill-coma recovery time of these beetles, suggesting that cold hardiness was reduced in its absence. These results suggest that the cold hardiness of O. communa may be partly regulated by MRLC-sqh. Our findings highlight the importance of motor proteins in mediating the cold response in insects.