Yang Zituo, Wong Sek Man, Yue Gen Hua
Department of Biological Sciences, National University of Singapore, 14 Science Drive, Singapore, 117543, Singapore.
Temasek Life Sciences Laboratory, National University of Singapore, 1 Research Link, Singapore, 117604, Singapore.
Mar Biotechnol (NY). 2021 Dec;23(6):854-869. doi: 10.1007/s10126-021-10068-x. Epub 2021 Nov 4.
Viral nervous necrosis (VNN) disease caused by the nervous necrosis virus (NNV) is a major disease, leading to a huge economic loss in aquaculture. Previous GWAS and QTL mapping have identified a major QTL for NNV resistance in linkage group 20 in Asian seabass. However, no causative gene for NNV resistance has been identified. In this study, RNA-seq from brains of Asian seabass fingerlings challenged with NNV at four time points (5, 10, 15 and 20 days post-challenge) identified 1228, 245, 189 and 134 DEGs, respectively. Eight DEGs, including rrm1, were located in the major QTL for NNV resistance. An association study in 445 survived and 608 dead fingerlings after NNV challenge revealed that the SNP in rrm1 were significantly associated with NNV resistance. Therefore, rrm1 was selected for functional analysis, as a candidate gene for NNV resistance. The expression of rrm1 was significantly increased in the gill, liver, spleen and muscle, and was suppressed in the brain, gut and skin after NNV challenge. The rrm1 protein was localized in the nuclear membrane. Over-expression of rrm1 significantly decreased viral RNA and titer in NNV-infected Asian seabass cells, whereas knock-down of rrm1 significantly increased viral RNA and titer in NNV-infected Asian seabass cells. The rrm1 knockout heterozygous zebrafish was more susceptible to NNV infection. Our study suggests that rrm1 is one of the causative genes for NNV resistance and the SNP in the gene may be applied for accelerating genetic improvement for NNV resistance.
由神经坏死病毒（NNV）引起的病毒性神经坏死（VNN）疾病是一种主要疾病，给水产养殖业造成了巨大的经济损失。先前的全基因组关联研究（GWAS）和数量性状基因座（QTL）定位已在尖吻鲈的20号连锁群中确定了一个抗NNV的主要QTL。然而，尚未确定抗NNV的致病基因。在本研究中，对在四个时间点（攻毒后5、10、15和20天）用NNV攻毒的尖吻鲈幼鱼的大脑进行RNA测序，分别鉴定出1228、245、189和134个差异表达基因（DEG）。包括rrm1在内的8个DEG位于抗NNV的主要QTL中。对445条在NNV攻毒后存活的和608条死亡的幼鱼进行的关联研究表明，rrm1中的单核苷酸多态性（SNP）与抗NNV显著相关。因此，选择rrm1进行功能分析，作为抗NNV的候选基因。在NNV攻毒后，rrm1在鳃、肝脏、脾脏和肌肉中的表达显著增加，而在大脑、肠道和皮肤中受到抑制。rrm1蛋白定位于核膜。rrm1的过表达显著降低了NNV感染的尖吻鲈细胞中的病毒RNA和滴度，而rrm1的敲低则显著增加了NNV感染的尖吻鲈细胞中的病毒RNA和滴度。rrm1基因敲除的杂合斑马鱼对NNV感染更敏感。我们的研究表明，rrm1是抗NNV的致病基因之一，该基因中的SNP可用于加速抗NNV的遗传改良。
