• Fisheries and Aquatic Sciences
• /
• 제13권4호
• /
• pp.263-271
• /
• 2010

G protein-coupled receptors (GPCR) constitute the largest superfamily of cell membrane receptors, mediating diverse signal-transduction pathways. The melanocortin 4 receptor (MC4R) has been of interest for its physiological role and size, one of the smallest among the GPCRs, which makes it a good model system for the structural study of GPCRs. To study the molecular structure and tissue-specific expression of MC4R in olive flounder (Paralichthys olivaceus), the full-length MC4R gene was obtained using PCR amplification of genomic DNA as well as cDNA synthesis. Sequence analysis of the gene indicates that 978 bp of the MC4R gene encodes 325 amino acids without introns. Sequence alignment with the MC4Rs from other fish shows the highest degree of identity (96%) between Paralichthys olivaceous and Verasper moseri, followed by Takifugu rubripes and Tetraodon nigroviridis (89%). RNA was isolated from various tissues to examine the tissue distribution of MC4R by using RT-PCR. The results showed major expression of MC4R in the liver, brain, and eye, which is consistent with the expression pattern in other fish belonging to the order Pleuronectiformes.

• Journal of Animal Science and Technology
• /
• 제49권4호
• /
• pp.437-442
• /
• 2007

본 연구는 축산과학원에서 보유중인 듀록 품종내에서 MC4R 유전자의 SNP 발현 특성과 육종가에 의한 선발 후 MC4R 유전자 빈도의 변화 및 경제형질에 대한 유전자형간 육종가의 차이를 구명하고자 수행하였다. 1999년부터 2005년까지 검정된 검정성적을 바탕으로 일당증체량, 등지방두께, 90kg 도달일령 및 사료요구율에 대하여 유전력과 유전상관 및 육종가를 추정하였으며, 2003년과 2004년에 출생한 660두에 대한 혈액을 채취하여 MC4R 유전자에 대한 유전자형 및 대립유전자 분석을 실시하였다. 또한 육종가에 의한 선발 후 유전자의 빈도변화를 세대별 그리고 선발군 및 도태군에 대하여 분석하였으며, 육종가의 분석결과를 이용하여 MC4R의 유전자형 효과를 보았다. 분석결과 육종가를 근거하여 선발한 MC4R 유전자형은 세대당 그리고 선발군과 도태군에서 차이를 보였으며, 각 형질별 육종가의 분산분석결과 사료요구율을 제외한 기타 경제형질에서 유의적으로 MC4R 유전자의 효과가 있는 것으로 나타났다. MC4R 유전자의 효과에 대한 보고는 상이한 부분도 있지만 자체 축군에 대한 다형성 분석 및 경제형질과의 연관성 분석에 의하여 축군 및 개량하고자 하는 경제형질에 따라 표지 유전인자로 활용하여 선발반응 및 정확도를 높일 수 있을 것으로 사료된다.

• Genes and Genomics
• /
• 제40권11호
• /
• pp.1119-1125
• /
• 2018

Melanocortin 4 receptor: (MC4R) and Myostatin (MSTN) are two important growth trait-related genes in animals. In this study, we showed that two SNPs, MC4R-719A>G and MSTN-519C>T, found in the promoters of the MC4R and MSTN genes, respectively, are both associated with growth traits in Spinibarbus hollandi. Furthermore, we observed that there were significant associations between the expression levels of the MC4R and MSTN genes and these two growth trait-related SNPs. The expression level of MC4R gene in brain was lower in GG genotype fish with extremely high growth performance than that in AA genotype fish with extremely low growth performance. Expression level of the MSTN gene in muscle was lower in TT genotype fish with extremely high growth performance than that in CC and CT genotype fish with lower growth performance. The results indicated that these SNPs located in the promoters of MC4R and MSTN are associated with growth-related traits through modification of gene expression levels. The MSTN and MC4R SNPs may have useful application in effective marker-assisted selection aimed to increase output in S. hollandi.

• 생명과학회지
• /
• 제15권6호
• /
• pp.968-971
• /
• 2005

본 연구는 Duroc, Landrace, Berkshire, Yorkshire를 기초 축으로 이용한 1003두에 대해 MC4R유전자의 PCR-RFLP를 이용하여 그 다형성을 조사하고 돼지의 일당증체량, 등지방 두께, 사료 요구율, 정육율과 그 유전자형 간의 연관성을 규명하고자 실시하였다. MC4R유전자에 대해 PCR-RFLP를 이용하여 226bp산물을 증폭한후 Taq I 체한효소로 사용하였다. 얻어진 MC4R gene의 유전자 빈도는 품종별로 다르게 나타났다. 통계적 분석을 통하여 각 유전자형에 대한 경제형질과 관련성을 분석한 결과 일당 증체량과 사료요구량은 NN 유전자형을 가진 개체들이 DN이나 DD유전자형을 가진 개체들에 비해 유의적으로 우수한 능력을 보였다(P < 0.05). 하지만 D 대립유전자는 높은 정육율과 낮은 등지방두께에 연관성이 있음을 관찰하였다. 따라서 돼지의 성장과 정육율과 관련된 선발력을 높이기 위해서 MC4R유전자의 다형성분석에서 검증된 PCR marker를 우량돼지육종 계획에 있어 분자생물학적 선발 marker로 사용할 수 있을 것으로 사료된다.

• 한국자기공명학회논문지
• /
• 제14권2호
• /
• pp.88-104
• /
• 2010

Human melanocortin-4 receptor (hMC4R) has a critical role in part of energy homeostasis, and their heterozygous mutations related in genetic cause of severe human obesity. In order to study the structure and function of these membrane proteins, it is important to prepare the samples. However, the preparation of transmembrane peptide is seriously difficult and time-consuming. Overexpression and purification of membrane proteins was reported to be difficult due to their innate insoluble and toxic properties. Among the many difficulties, the most important is the difficulty in obtaining sufficient quantities of purified protein. Recently, we succeed to produce large amounts of the second transmembrane domain from the wild-type hMC4R (wt-TM2) and D90N mutant hMC4R (m-TM2) and proposed the structural difference of them in membrane-like environments. In this paper, we demonstrate the optimization procedures to express and purify wt-TM2 or m-TM2 peptides, and solution NMR studies in different detergents to get high-resolution spectra were also described.

• Asian-Australasian Journal of Animal Sciences
• /
• 제19권6호
• /
• pp.763-768
• /
• 2006

Genetic and pharmacological studies in mice have demonstrated a complementary role for the melanocortin 4 receptor (MC4R) in the control of food intake, energy balance and body weight. This study was designed to investigate the associations of a MC4R gene polymorphism on chicken growth and body composition traits in broiler lines divergently selected for abdominal fat. A SNP (G54C) was found in CDS region of chicken MC4R gene. The analysis of the least squares and variance revealed a significant association between the G54C SNP and BW, CW and SL at 7 wk of age, and there were significant differences in different genotypes (p<0.05). The results from protein secondary structure prediction and tertiary structure prediction showed that it appeared a helix in $13^{th}$ amino acid and two strands at $14^{th}$ and $15^{th}$ amino acid in mutant protein, respectively. It maybe induce the change of the activity or function of MC4R gene in poultry.

• 한국자기공명학회논문지
• /
• 제16권1호
• /
• pp.34-45
• /
• 2012

Melanocortin-4 receptor (MC4R) subtype is associated with obese humans. Especially, in a patient with severe early-onset obesity, novel heterozygous mutation in the MC4R gene was detected, resulting in an exchange of aspartic acid to asparagine in $90^{th}$ amino acid residue located in the predicted second trans-membrane domain (TM2). Mutations in the melanocortin-4 receptor (MC4R) gene are the most frequent monogenic causes of severe obesity which have been described as heterozygous with loss of function. In order to compare structure difference between MC4R wild type (MC4R-TM2-wt) and mutant (MC4R-TM2-D90N), we designed both MC4R-TM2-wt and MC4R-TM2-D90N construct in pET 21b vector. In this study, we optimized high-yield purification procedure for recombinant TM2-D90N. Eluted recombinant protein was resolubilized under urea condition for thrombin cleavage reaction and we conducted the high-performance liquid chromatography (HPLC) with reverse phase column under 1% acetonitrile, 0.01% TFA buffer solution. The molecular size of purified target peptide was confirmed by Tricine-SDS page analysis. To characterize MC4R-TM2-D90N, we have performed $^{15}N$-isotope labeling of peptide using M9 media and purified labeled target peptide for hetero-nuclear NMR spectroscopy.

• 한국발생생물학회지:발생과생식
• /
• 제14권4호
• /
• pp.261-268
• /
• 2010

As in the O. mykiss electrophoretic profiles of RNA, the signals of each RNA sample from 9 individual tissues such as liver, muscle, brain, heart, pituitary gland, kidney, intestine, spleen and gill similar to positive control were obtained. The tissue distributions of the complimentary DNA (cDNA) of O. mykiss four genes were analyzed using quantitative real-time PCR with primer sets for tissue expression analysis. In this rainbow trout species, author obtained bands of various sizes, ranged from 700 bp to 1,400 bp. A dissociation curve was made at the end of each run to make sure that there was no non-specific amplification. Supplementarily, the Ct of each DNA was compared. The Ct values of vinculin with rainbow trout tissues were determined in a manner similar to those for agouti-related protein (AgRP) and melanocortin receptors (MC4R I and MC4R II). Further, obtained Cts for standard curve of each DNA were affected by specific product (vinculin, AgRP and MC4R II genes). After several experiments with four individual genes of rainbow trout, author estimated a variation ratio of the mean Ct value of the DNA extracted using the comparative CTt method was 37.27, and the standard deviation was 5.33. The correlation coefficient between the Ct values and the concentration of cDNA was -0.98, -0.99, -0.91 and -0.86, respectively (vinculin, AgRP, MC4R I and MC4R II genes). Since this correlation showed high linearity, the straight line obtained was used as a standard for the O. mykiss tissues reared in aquarium. A PCR efficiency of 100% is ideally achieved when the slopes are close to the theoretical value of -3.31. According to quantification method, the results of quantification are strongly affected by the DNA fragmentation. The size of most DNA fragments obtained from various tissues of rainbow trout used in the experiment was approximately 100 bp. According to the four slopes, an efficiency of nearly 100% was estimated for four genes detection methods. Additionally, further analysis with more individuals and primers will be required to fully establish optimization in rainbow trout.

• 생명과학회지
• /
• 제30권1호
• /
• pp.58-63
• /
• 2020

본 연구에서는 제주재래돼지와 듀록 품종 사이에서 생산된 F₂ 참조축군에서 melanocortin-4-receptor (MC4R) 유전자형과 지방산 조성간의 관련성을 연구하였다. 전체 290 여두의 F₂ 자손을 이용하여 14개의 지방산 조성을 측정하였다. Taq I PCR-RFLP 방법을 이용하여 MC4R c.1426A>G (p.Asp298Asn)의 단일염기다형성(single nucleotide polymorphism, SNP)을 확인하였다. MC4R 세 가지 유전자형(AA, AB, BB)이 모두 발견되었고, 그 빈도는 각각 0.299, 0.542, 0.159로 확인되었다. AA 유전자형을 가진 개체에서 palmitic acid (C16:0, p<0.05), stearic acid (C18:0, p<0.01), eicosenoic acid (C20:1n9, p<0.05), saturated fatty acid (SFA, p<0.01) 함량이 GG 유전자형을 가진 개체보다 더 높은 것으로 확인되었다. 반면에 GG 유전자형을 가진 개체는 linoleic acid (C18:2n6, p<0.001), linolenic acid (C18:3n3, p<0.001), linolenic acid (C18:3n6, p<0.001), arachidonic acid (C20:4n6, p<0.001)과 같은 불포화지방산(unsaturated fatty acid) 함량이 AA 유전자형을 가진 개체보다 더 높은 것을 확인하였다. 제주재래돼지와 듀록 품종 사이에서 생산된 F₂ 참조축군에서 MC4R GG 유전자형이 포화지방산은 낮추고, 불포화지방산은 높이는 것으로 확인되었다. 본 연구 결과 MC4R의 유전적 다형성이 듀록과 제주재래돼지 교배 프로그램에서 육질 향상과, 고기내의 지방산 함량을 조절할 수 있는 유전적 표지 인자로 활용될 수 있을 것이라 사료된다.

• 한국수정란이식학회지
• /
• 제28권3호
• /
• pp.215-222
• /
• 2013

Bovine coat color is decided by the melanocortin receptor 1 (MC1R) genotype mutation and melanogenesis. Specially, in the various cattle breeds, dominant black coat color is expressed by dominant genotype of $E^D$, red or brown is expressed in the frame shift mutation of recessive homozygous e by base pair deletion and wild type of $E^+$ is expressed in various coat colors. However, not very well known about the effected of MC1R genotype mutation on the coat color through family lines in KBC. Therefore, this study were to investigate effect of MC1R genotype mutation on the coat color, and to suggest mating breed system in accordance with of MC1R genotype for increased on brindle coat color appearance. Parents (sire 2 heads and dam 3 heads) and offspring (total : 54 heads) from crossbreeding in KBC family line with the MC1R genotype and phenotype records were selected as experimental animals. The relationship between melanocortin 1 receptor (MC1R) genotypes expression verified by PCR-RFLP, and brindle coat color appearance to the family line of the cross mating breed from MC1R genotype pattern was determined. As a result, 4MC1R genetic variations, $E^+/E^+$ (sire 1), $E^+/e$ (sire 2 and dam 3), $E^+/e$ with 4 bands of 174, 207 and 328 bp (dam 1) and $E^+/e$ with 3 bands of 174, 207, 328 and 535 bp (dam 2) from parents (sire and dam) of KBC. However, 3 genetic variations, e/e (24%), $E^+/E^+$ (22%) and $E^+/e$ (56%) were identified in offspring. Also, brindle coat color expressrated was the e/e with the 0%, $E^+/E^+$ with 67% and $E^+/e$ with 77% from MC1R genotype in offspring on the cross mating of KBC. Furthermore, when the sire had $E^+/e$ genotype and the dam had $E^+/E^+$ with the 3 bands or $E^+/e$ genotype, and both had whole body-brindle coat color, 62% of the offspring had whole body-brindle coat color. Therefore, the seresults, the mating system from MC1R genotype patterns of the sires ($E^+/e$) and dams ($E^+/E^+$ with the 3 bands or $E^+/e$) with brindle coat color may have the highest whole body-brindle coat color expression in their offspring.