Cloning and Expression Analysis of the ??Subunit of Porcine Prolyl 4-hydroxylase

Eun Seok Cho,Eun Jung Kwon,Won Youg Jung,Da Hye Park,Ki Hwa Chung,Kwang Keun Cho,Chul Wook Kim

doi:10.5713/ajas.2007.1655

Abstract

Prolyl 4-hydroxylase (P4H) plays a central role in collagen synthesis by catalyzing the hydroxylation of the proline residue in the X-Pro-Gly amino acid sequence, and controls the biosynthesis of collagen that influences overall meat quality. In order to verify expression level of the catalytic a subunit of P4H, a 2.7 kb clone of the α-subunit gene for P4H was selected from a cDNA library prepared from the muscular tissue of Sancheong berkshire pigs, and the whole gene sequence was determined. As expression level of the α-subunit of P4H differed between tissues of pigs, we intended to assess more precisely the level of α-subunit expression between tissues of Sancheong Berkshire pigs by using RT-PCR. Muscular and adipose tissues were taken from each pig grouped by growth stage (weighing 60, 80, and 110 kg) of Yorkshire and Sancheong Berkshire pigs, and the expression levels of the α-subunit of P4H were examined. Since there were significant differences in the expression level with respect to variation in growth stage (p<0.01), an attempt was made to identify any influences of pig species and tissue variation. The muscular and adipose tissues of pigs weighing 110 kg showed higher expression levels than pigs weighing 60 kg and 80 kg. In general, significantly higher expression levels were found in muscular than in adipose tissue. The expression levels in Sancheong Berkshire were significantly higher than in Yorkshire pigs (p<0.01 or p<0.05). Since expression level of the α-subunit of P4H affects the activity of P4H and is connected to the biosynthesis of collagen and increased collagen can improve meat texture, this finding may explain why meat quality of the Sancheong Berkshire pig is acclaimed in Korea. Given the higher expression levels of the α-subunit gene in adipose than in muscular tissue, and also in the heavier pigs, more intensive studies are required to assess the correlation between expression level of the α-subunit gene and overall meat quality.

Highlights

Prolyl 4-hydroxylase catalyzes the hydroxylation of proline residues in repeating X-Pro-Gly triplets, forming the 4-hydroxyproline required for the correct folding of newly synthesized collagen (Kivirikko and Myllyharju, 1998; Kivirikko and Pihlajaniemi, 1998; Myllyharju, 2003)
The results showed a higher level of expression in the adipose tissue of pigs weighing 110 kg, which suggests that level of expression of the α-subunit of Prolyl 4-hydroxylase (P4H) is closely associated with the growth stage of pigs, regardless of choice of tissues
The highest expression level appeared in adipose tissue of pigs weighing 110 kg, whereas much was seen in pigs weighing 80 kg which was similar to that lower level of expression was observed in muscular tissue, of Sancheong Berkshire pigs (Figure 3)

Summary

INTRODUCTION

Prolyl 4-hydroxylase catalyzes the hydroxylation of proline residues in repeating X-Pro-Gly triplets, forming the 4-hydroxyproline required for the correct folding of newly synthesized collagen (Kivirikko and Myllyharju, 1998; Kivirikko and Pihlajaniemi, 1998; Myllyharju, 2003). It has been reported that concentration of the α-subunit of P4H was correlated with cell density and had a positive effect upon collagen formation (Rowe and Schwarz, 1983; Kao et al, 1985). Regulation of the amount of active prolyl 4-hydroxylase tetramer appears to be linked to synthesis of the α-subunit (Kivirkko et al, 1992). It was presumed that the expression of the α-subunit of P4H involved in collagen biosynthesis might be related to growth phase and/or meat quality of pigs. It is known that collagen is a key factor for the elasticity of skin and muscle of mammals, and its adhesive property is responsible for linking of cells, which is related to the firmness of pig muscle. We attempted to determine differences in expression level of the α-subunit of P4H between each tissue of the pig in order to clarify whether differential expression could be correlated with tissue specificity and sub-speciation of pigs

MATERIALS AND METHODS

RESULTS

DISCUSSION