• BMB Reports
• /
• v.40 no.3
• /
• pp.432-438
• /
• 2007

V(D)J recombination, a site-specific gene rearrangement process occurring during the lymphocyte development, begins with DNA double strand breaks by two recombination activating gene products (RAG1/2) and finishes with the repair process by several proteins including DNA-dependent protein kinase (DNA-PK). In this report, we found that RAG2 was specifically phosphorylated by DNA-PK at the $365^{th}$ serine residue, and this phosphorylated RAG2 affected the V(D)J recombination activity in cells in the GFP expression-based assay. While the V(D)J recombination activity between wild-type RAG2 and mutant S365A RAG2 in the assay using a signal joint substrate was undistinguishable in DNA-PK deficient cells (M059J), the activity with wild-type RAG2 was largely increased in DNA-PK proficient cells (M059K) in comparison with mutant RAG2, suggesting that RAG2 phosphorylation by DNA-PK plays a crucial role in the signal joint formation during V(D)J recombination.

• Korean Journal of Fisheries and Aquatic Sciences
• /
• v.57 no.3
• /
• pp.239-252
• /
• 2024

Recombination activating genes (RAGs) play a crucial role in initiating V(D)J recombination, which is essential for developing adaptive immunity in vertebrates. In this study, we cloned and characterized RAG1/2 cDNA from the marine medaka Oryzias dancena (OdRAG1/2) and investigated their mRNA expression patterns during ontogenetic developmental stages. The OdRAG1 and OdRAG2 cDNA contained open reading frames (ORFs) encoding proteins containing 1,078 and 531 amino acids, respectively. Multiple sequence alignment and phylogenetic analysis revealed that OdRAG1 and OdRAG2 are highly conserved with their corresponding orthologs, featuring distinct core and non-core regions. Notably, expression analysis showed that, in contrast to other fish RAGs studied, OdRAG1/2 expression peaked at 0 days post-hatching (DPH). Additionally, for the expression of T and B cell differentiation markers, CD3γ and CD20, also peaked at 0 DPH. Collectively, adaptive immunity in O. dancena potentially begins during embryonic development, which is critical for V(D)J recombination and essential immune component development, suggesting the early ontogenetic stage interactions between innate and adaptive immunity.

• Korean Journal of Ichthyology
• /
• v.22 no.4
• /
• pp.273-278
• /
• 2010

Partial nucleotide sequences of nuclear protein-coding recombination activating gene 1 (RAG1) gene of two Abbottina and five Microphysogobio species residing in Korea were analyzed to elucidate the molecular phylogenetic position of A. springeri Banarescu and Nalbant. In RAG1 tree A. rivularis was clearly separated from the monophyletic lineage composed of A. springeri, Biwia zezera and Microphysogobio species. Within this lineage B. zezera showed sister-group relationship to the monophyletic group composed of A. springeri and five Microphysogobio species. Thus, our phylogenetic tree revealed the polyphyletic nature of two Abbottina species from Korea, which result is well congruent with the previous phyletic assumption based on osteological features. The current classification of Abbottina and Microphysogobio based on morphological criteria, such as the presence or absence of papillae on lips and size of swim bladder with or without encapsulation, does not reflect their true evolutionary history.

• BMB Reports
• /
• v.36 no.2
• /
• pp.201-206
• /
• 2003

Two lymphoid-specific proteins, RAG1 and RAG2, are required for the initiation of the V(D)J recombination in vitro. The V(D)J cleavage that is mediated by RAG proteins at the border between the coding and signal sequences results in the production of a hairpin at the coding end and a double-stranded break at the signal end. Two hairpin coding ends are re-opened, modified, and sealed; whereas, the signal ends are directly ligated. Here I report that only RAG1 can carry out a distinct endonucleolytic activity in vitro using an oligonucleotide substrate that is tethered by a short single-stranded DNA. The purified RAG1 protein alone formed a nick at the near position to the recombination signal sequence. This endonucleolytic activity was eliminated by immunoprecipitation using the RAG1-specific antibody, and required the 3'-hydroxy group. All of the RAG1 mutants that were incapable of the nick and hairpin formation in the V(D)J cleavage analysis also showed this new endonucleolytic activity. This suggests that the nicking activity that was observed might be functionally different from the nick formation in the V(D)J cleavage.

• Korean Journal of Ichthyology
• /
• v.21 no.4
• /
• pp.287-290
• /
• 2009

A natural hybrid between the striped spine loach Cobitis tetralineata and the king spine loach Iksookimia longicorpa was genetically identified by sequence analyses of nuclear recombination activating gene 1 (RAG1) and mitochondrial cytochrome c oxidase I (COI) genes. Out of 850 base positions of RAG1, a total of 23 nucleotide substitutions were detected between the two parental species, whereas the electropherogram of the natural hybrid displayed double peaks at all of the 23 positions, which reflects their simple Mendelian inheritance pattern. Meanwhile, comparison of partial sequences of mitochondrial genes (COI in this study), which are well characterized by the maternal inheritance pattern, revealed that the maternal species of the hybrid was C. tetralineata because of their 100% sequence identity.

• Korean Journal of Ichthyology
• /
• v.27 no.3
• /
• pp.199-204
• /
• 2015

A natural hybrid between Acheilognathus signifer and A. lanceolatus was collected in their sympatric area, the Gimhwanamdaecheon of Hangang (River), Korea. Morphological characters, mitochondrial cytochrome b gene (cyt b), and recombination-activating gene 1 (RAG-1) were investigated to confirm the natural hybrid origin. As a result of morphological characters, the natural hybrid was appeared to have intermediate characters between two parental species in three characters; the band of dorsal fin, the color patterns of anal fin membrane, and the body color. In analysis of cyt b, it was revealed that the maternal species of the natural hybrid was appeared to be A. signifer due to their 99.9% sequence identity. Also, in analysis of RAG-1, an electropherogram of the hybrid individual displayed double peaks, strongly indicating its hybrid state.

• Korean Journal of Ecology and Environment
• /
• v.53 no.3
• /
• pp.275-285
• /
• 2020

Two specimens presumed to be hybrids of Rhodeus pseudosericeus and R. notatus were collected from the Sangcheon Stream of the Han River, Korea. The body color of natural hybrid individuals was yellowish brown, showing the intermediate characteristics of R. pseudosericeus and R. notatus, but overall, the characteristics of R. notatus were similar. Meristic and morphometric characters, the number of dorsal fin rays, anal fin rays and longitudinal row scales showed a hybrid index (HI) of 0, indicating the characters of R. notatus. The hybrid index of predorsal length (HI=74.6), preanal length (HI=75.3), and preventral length (HI=77.6) were similar to the characters of the R. pseudosericeus. Also four characters were appeared to have intermediate characters between R. pseudosericeus and R. notatus; number of gill rakers (HI=55.3), body depth (HI=67.9), snout length (HI=43.4), and inter orbital width (HI=44.8). The rest of 14 characters deviated between 0 and 100, showing unique characteristics of hybrid individuals. Recombination activating gene 1 (RAG1) analysis, hybrids were identified as natural hybrids due to the overlapping peaks of their parent species. Also, as a result of analysis using the cytochrome b gene (COB), one individual was derived from R. pseudosericeus, while the other was derived from R. notatus.

• Korean Journal of Ichthyology
• /
• v.26 no.1
• /
• pp.70-73
• /
• 2014

Interspecific hybrids between tiger grouper Epinephelus fuscoguttatus and giant grouper E. lanceolatus were genetically identified based on the partial sequence analysis of mitochondrial cytochrome c oxidase I (COX I) gene and nuclear recombination activating gene 2 (RAG 2) gene. Out of 585 base positions of RAG 2, a total of five nucleotide substitutions were detected between the two parental species (E. fuscoguttatus and E. lanceolatus). The hybrids had two distinct types of RAG 2 sequences corresponding to those of both parental species. Mitochondrial COX I gene sequencing showed that hybrids had sequences identical to E. fuscoguttatus. Molecular data clearly demonstrate that hybridization does occur between E. fuscoguttatus and E. lanceolatus, but with E. fuscoguttatus as the maternal parent.

• Journal of Aquaculture
• /
• v.22 no.1
• /
• pp.79-82
• /
• 2009

Reciprocal interspecific hybrids between two bitterling species Rhodeus uyekii (RU) and R. notatus (RN) were genetically identified based on the partial sequence analysis of recombination activating gene-1 (RAG-1) gene. Out of 863 bp positions analyzed, 13 nucleotide substitutions were detected between the two parental species (RU and RN genotypes). Both the induced hybrids (RU female$\times$RN male; UN genotype) and their reciprocal counterparts (RN female$\times$RU male; NU genotype) displayed the double peaks (or polymorphism) of sequence chromatograms at the 13 diagnostic positions, indicating that those hybrids were actual karyogamy derived from the two parental haploid genomes. However, it was not possible to distinguish between the reciprocal interspecific hybrids.

• Korean Journal of Ichthyology
• /
• v.29 no.1
• /
• pp.1-12
• /
• 2017

A natural hybrid of interspecific between the Coreoleuciscus splendidus and C. aeruginos (Cypriniformes: Cyprinidae) was captured in the Hwang-ji Stream, a tributary of the Nakdong River basin in Korea. An interspecific hybrid between C. splendidus and C. aeruginos was genetically identified based on morphological characteristics and the sequence analysis of nuclear recombination activating gene 1 (RAG1) gene (1,334 bp) and mitochondrial cytochrome c oxidase subunit 1 (CO1) gene (1,551 bp). As a result of morphological variations, the natural hybrid appeared to have an intermediate character between two parental species (C. splendidus and C. aeruginos) in three variations of black array (s) on dorsal, caudal and anal fin rays. Phylogenetic analysis inferred from RAG1 and CO1 sequence data revealed that Coreoleuciscus populations from Hwang-ji stream consist of two pure Coreoleuciscus species and a hybrid individual group. The individuals were clearly identified the cross and reciprocal hybrid by CO1 gene analysis. In RAG1 gene, 13 nucleotide variation loci were detected and the hybrid individuals displayed the double peaks of sequence chromatograms at the 9 diagnostic positions. In this study, molecular data and morphological variations were clearly demonstrated that hybridization did occur between C. splendidus and C. aeruginos. However, F2 hybrid generation and reproductive capacity of F1 hybrid individuals were not demonstrated.