Characterization of Histone Gene Expression in Sevenband Grouper, Hyporthodus septemfasciatus against Nervous Necrosis Virus Infection

Abstract

Sevenband grouper (Hyporthodus septemfasciatus) is a high-quality fish that is one of the most popular seafood species for consumption. A group of deaths occurred in fish farms as a result of an infection by the Nervous Necrosis Virus (NNV), that causes viral nervous necrosis (VNN) disease. Studies of other fish species revealed that histone proteins are involved in innate immune responses during pathogen invasion as well as DNA packing. In this study, we characterized the histone ORF genes (H1, H2A, H2A.V, H3.3-1, H3.3-2) of sevenband groupers and analyzed histone gene expression in NNV-infected sevenband groupers. The histone gene amino acid sequences of sevenband grouper were similar among fishes and even with mammals and were an evolutionarily conservative. High gene expression of histone genes was observed in the tissues of sevenband grouper infected by NNV. Quantitative real time PCR analysis of cDNA showed that H1 had a high gene expression level in brain and eye on day 2 after infection, and H2A.V had a high gene expression level in brain and blood after NNV-infected. On day 2 after infection, H3.3-1 showed high gene expression in the liver and blood, and H3.3-2 showed the highest gene expression in the brain and blood. An increase in the amount of extracellular histone protein in the blood has been identified as a biomarker for vascular function in human, an increase in gene expression level of histone proteins in the blood of sevenband grouper implied that it is reasonable to assume that the histone gene is involved in the innate immune response in fish. More research is required to understand Histone's immune response at the protein level or in aquatic animals.

능성어는 고급 양식어종으로 식용으로 인기있는 해산어종 중의 하나이다. Viral nervous necrosis(VNN) 질병을 일으키는 Nervous necrosis virus(NNV) 감염으로 인해 양식장에 집단 폐사를 일으켜 많은 피해를 입히고 있다. 최근 histone protein이 DNA packing 뿐 아니라 선천적 면역 반응에 관여한다는 연구들이 보고되어, 능성어의 histone ORF genes(H1, H2A, H2A.V, H3.3-1, H3.3-2)들을 확인하고, NNV에 감염된 능성어의 histone genes의 조직 별 발현 양을 분석하였다. Histone gene들은 각 종 간 amino acid 서열이 유사하여, 진화적으로 보존적인 서열을 가지고 있음을 확인하였다. NNV에 감염된 능성어의 조직 별 발현 양 분석 결과 감염 후 2일차에 H1은 brain, eye에서 높은 유전자 발현양을 확인하였으며, H2A.V는 brain, blood에서 높은 유전자 발현양을 확인하였다. H3.3-1은 감염 후 2일차에 liver, blood에서 높은 유전자 발현양을 확인하였고, H3.3-2 유전자에서 brain, blood에서 유전자 발현양이 가장 높음을 확인하였다. 혈액 내 세포 외 histone protein 양의 증가는 혈관 기능 이상의 바이오마커로 알려져 있어, histone 유전자는 선천적인 면역반응에 관여함을 추측할 수 있다. Histone의 면역반응을 단백질 수준에서 설명하기 위해서는 추가적인 연구가 필요하다.