RESUMEN
The Rab protein family belongs to a superfamily of ras-like GTP-binding proteins. Rab proteins regulate many steps of membrane trafficking. In this study, three Rab family members, Rab5B, Rab6A, and Rab7, designated LvRab5B, LvRab6A, and LvRab7, were cloned from Litopenaeus vannamei. The full-length cDNA sequences of LvRab5B, LvRab6A, and LvRab7 were 1383, 873, and 767 nucleotides in length and they encoded proteins of 211, 212, and 205 amino acids, respectively. Using qRT-PCR, the mRNA expression levels of the three proteins were determined in the hepatopancreas of L. vannamei at different stages after infectious hypodermal and hematopoietic necrosis virus and white spot syndrome virus challenge. The results indicated that the mRNA expression levels of LvRab5B, LvRab6A, and LvRab7 were all significantly up-regulated after virus injection, suggesting that these genes may play essential roles in the immune response to viral infection in shrimp.
Asunto(s)
Regulación de la Expresión Génica , Penaeidae/genética , Proteínas de Unión al GTP rab/genética , Proteínas de Unión al GTP rab5/genética , Secuencia de Aminoácidos , Animales , Western Blotting , Clonación Molecular , Densovirinae , Perfilación de la Expresión Génica , Datos de Secuencia Molecular , Filogenia , Reacción en Cadena de la Polimerasa , ARN Mensajero/genética , ARN Mensajero/metabolismo , Proteínas Recombinantes/metabolismo , Alineación de Secuencia , Análisis de Secuencia de ADN , Virus del Síndrome de la Mancha Blanca 1 , Proteínas de Unión al GTP rab/química , Proteínas de Unión al GTP rab/metabolismo , Proteínas de Unión al GTP rab5/química , Proteínas de Unión al GTP rab5/metabolismo , Proteínas de Unión a GTP rab7RESUMEN
MicroRNAs (miRNAs) are known to play an important role in regulating both adaptive and innate immunity. Pacific white shrimp (Litopenaeus vannamei) is the most widely farmed crustacean species in the world. However, little is known about the role miRNAs play in shrimp immunity. To understand the impact of viral infection on miRNA expression in shrimp, we used high-throughput sequencing technology to sequence two small RNA libraries prepared from L. vannamei under normal and white spot syndrome virus (WSSV) challenged conditions. Approximately 19,312,189 and 39,763,551 raw reads corresponding to 17,414,787 and 28,633,379 high-quality mappable reads were obtained from the two libraries, respectively. Twelve conserved miRNAs and one novel miRNA that were highly expressed (>100 RPM) in L. vannamei were identified. Of the identified miRNAs, 8 were differentially expressed in response to the virus infection, of which 1 was upregulated and 7 were downregulated. The prediction of miRNA targets showed that the target genes of the differentially expressed miRNAs were related to immunity, apoptosis, and development functions. Our study provides the first characterization of L. vannamei miRNAs in response to WSSV infection, which will help to reveal the roles of miRNAs in the antiviral mechanisms of shrimp.
Asunto(s)
Inmunidad Innata/genética , MicroARNs/genética , Penaeidae/genética , Virus del Síndrome de la Mancha Blanca 1/genética , Animales , Regulación de la Expresión Génica , MicroARNs/biosíntesis , Penaeidae/virología , Virus del Síndrome de la Mancha Blanca 1/patogenicidadRESUMEN
The macrophage migration inhibitory factor (MIF) is an important proinflammatory cytokine that mediates both innate and adaptive immune responses. In this study, we identified a homolog of MIF in the Pacific white shrimp Litopenaeus vannamei. The MIF cDNA contained a 363-bp open reading frame encoding a 120-amino acid protein with a calculated molecular mass of 13.442 kDa and a theoretical isoelectric point of 6.57. The L. vannamei MIF shared high amino acid identity with MIFs of other invertebrates. Tissue distribution analysis by quantitative real-time polymerase chain reaction (qRT-PCR) revealed that the L. vannamei MIF was abundantly expressed in the blood, heart, and hepatopancreas, was moderately expressed in the gill, and was weakly expressed in the muscle and intestine. Furthermore, in order to gain a basic understanding of the role of MIF in the shrimp immune response against viral infection, its mRNA expression was determined in the hepatopancreas of L. vannamei at different stages after white spot syndrome virus (WSSV) challenge using qRT-PCR. The result indicated that the expression of MIF was significantly upregulated after WSSV injection, suggesting that MIF may be involved in the response to viral infection in shrimp.
Asunto(s)
Factores Inhibidores de la Migración de Macrófagos/genética , Penaeidae/genética , Secuencia de Aminoácidos , Animales , Factores Inhibidores de la Migración de Macrófagos/química , Factores Inhibidores de la Migración de Macrófagos/metabolismo , Datos de Secuencia Molecular , Especificidad de Órganos , Penaeidae/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismoRESUMEN
Ryanodine receptor/calcium release channel is a large protein that plays an essential role in muscle contraction; mutations in the ryanodine receptor gene affect sensitivity to stress. As a first step towards investigating the relationship between the ryanodine receptor and shrimp cramped muscle syndrome, we cloned, partially sequenced, and examined single-nucleotide polymorphisms (SNPs) of the ryanodine receptor gene of the Pacific white shrimp (Litopenaeus vannamei). The nucleotide sequence of a 15.06-kb L. vannamei genomic DNA segment containing a partial ryanodine receptor gene sequence was determined (deposited in GenBank nucleotide database: HM367069). Direct sequencing of PCR-amplified ryanodine receptor exons with their intron-flanking regions in 10 cramped muscle syndrome shrimp and 10 healthy shrimp, revealed seven SNPs. Five of them (1713A/G, 1749T/C, 1755T/C, 3965G/A, and 8737C/T) are located in exons; however, they appear to be neutral (synonymous), since they do not alter the encoded amino acid. The other SNPs (1553C/T and 13337A/G) are in introns. The SNPs identified in the ryanodine receptor gene could be useful for association studies aimed at determining the physiological role of the ryanodine receptor in cramped muscle syndrome of shrimp.