Promoter polymorphisms in genes involved in porcine myogenesis influence their transcriptional activity.
BACKGROUND
Success of meat production and selection for improvement of meat quality is among the primary aims in animal production. Meat quality traits are economically important in swine; however, the underlying genetic nature is very complex. Therefore, an improved pork production strongly depends on identifying and studying how genetic variations contribute to modulate gene expression. Promoters are key regions in gene modulation as they harbour several binding motifs to transcription regulatory factors. Therefore, polymorphisms in these regions are likely to deeply affect RNA levels and consequently protein synthesis. In this study, we report the identification of single nucleotide polymorphisms (SNPs) in promoter regions of candidate genes involved in development, cellular differentiation and muscle growth in Sus scrofa. We identified SNPs in the promoter regions of genes belonging to the Myogenic Regulatory Factors (MRF) gene family (the Myogenic Differentiation gene, MYOD1) and to Growth and Differentiation Factors (GDF) gene family (Myostatin gene, MSTN, GDF8), in Casertana and Large White breeds. The purpose of this study was to investigate if polymorphisms in the promoters could affect the transcriptional activity of these genes. With this aim, we evaluated in vitro the functional activity of the luciferase reporter gene luc2 activity, driven by two constructs carrying different promoter haplotypes.


RESULTS
We tested the effects of the G302A (U12574) transition on the promoter efficiency in MYOD1 gene. We ascertained a difference in transcription efficiency for the two variants. A stronger activity of the A-carrying construct is more evident in C2C12. The luciferase expression driven by the MYOD1-A allelic variant displayed a 3.8-fold increased transcriptional activity. We investigated the activity of two haplotype variants (AY527152) in the promoter of GDF8 gene. The haploptype-1 (A435-A447-A879) up-regulated the expression of the reporter gene by a two-fold increase, and hence presumably of the GDF8 gene, in both CHO and C2C12 cultured cells.


CONCLUSIONS
In vitro the MYOD1-A allelic variant could up-regulate the expression of MYOD1 gene. Additionally, we could assess a different response of in vitro gene expression according to cell type used to transfect constructs, suggesting that MyoD activation is regulated by mechanisms that are specific of myoblasts.
Department for Innovation in Biological, Agro-food and Forest systems, University of Tuscia, Viterbo, 01100, Italy. bongiorni@unitus.it.
Silvia
S
Bongiorni
Department of Ecological and Biological Sciences, University of Tuscia, Viterbo, 01100, Italy. francesca.tilesi@unitus.it.
Francesca
F
Tilesi
Department for Innovation in Biological, Agro-food and Forest systems, University of Tuscia, Viterbo, 01100, Italy. silvia.bic@hotmail.it.
Silvia
S
Bicorgna
Department for Innovation in Biological, Agro-food and Forest systems, University of Tuscia, Viterbo, 01100, Italy. francesca_iacoponi@yahoo.it.
Francesca
F
Iacoponi
Department of Ecological and Biological Sciences, University of Tuscia, Viterbo, 01100, Italy. willems@unitus.it.
Daniela
D
Willems
Department for Innovation in Biological, Agro-food and Forest systems, University of Tuscia, Viterbo, 01100, Italy. maria.gargani@unitus.it.
Maria
M
Gargani
Department of Agricultural, Environmental and Food Sciences, University of Molise, Campobasso, 86100, Italy. dandrea@unimol.it.
MariaSilvia
M
D'Andrea
Department of Agricultural, Environmental and Food Sciences, University of Molise, Campobasso, 86100, Italy. Pilla@unimol.it.
Fabio
F
Pilla
Department for Innovation in Biological, Agro-food and Forest systems, University of Tuscia, Viterbo, 01100, Italy. alessio@unitus.it.
Alessio
A
Valentini
D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D063427:Food Quality; D005787:Gene Frequency; D008460:Meat; D051379:Mice; D024510:Muscle Development; D018482:Muscle, Skeletal / Q000254:growth & development / Q000378:metabolism; D017570:MyoD Protein / Q000235:genetics* / Q000378:metabolism; D055435:Myostatin / Q000235:genetics* / Q000378:metabolism; D010641:Phenotype; D020641:Polymorphism, Single Nucleotide; D011401:Promoter Regions, Genetic; D017422:Sequence Analysis, DNA; D034421:Sus scrofa / Q000235:genetics / Q000254:growth & development; D014158:Transcription, Genetic; D015533:Transcriptional Activation
D017570:MyoD Protein; C057363:MyoD1 myogenic differentiation protein; D055435:Myostatin