Translational efficiency of casein transcripts in the mammary tissue of lactating ruminants

Abstract

-Caseins are essentially concentrated in the colloidal fraction of ruminant milks as highly hydrated and mineralized spherical particles, termed casein micelles. They form a group of four peptide chains (α s1 , β, α s2 and κ), encoded by four structural genes (CSN1S1, CSN2, CSN1S2 and CSN3, respectively) of which the expression is regulated by lactogenic hormones. These phosphoproteins are synthesized, essentially during lactation, in the mammary epithelial cells and we show, for the first time, that their regulation is also controlled at the translational level. Apparently, the four casein messenger are not translated with the same efficiency. Specific amplification systems have been developed and optimized to quantify, by real time quantitative PCR (qPCR), transcripts encoding the four caseins starting from total RNA extracted from mammary tissues taken on goats (n = 4), ewes (n = 3) and cows (n = 3), in lactation. The relative proportions of each specific messenger (% of casein mRNA) were compared to the relative amounts of the corresponding caseins (% of whole casein) in milks sampled from the same animals, determined after fractionation by reverse phase HPLC and integration of the corresponding peak areas. From qPCR data, the four casein transcripts appeared to be present approximately at the same level of abundance (ca. 25%, except for defective genotypes at the CSN1S1 locus, in the goat) whereas the amounts of the corresponding proteins in milk were ranging between 9 and 38% of the whole casein fraction. A comparison of specific translational efficiencies (% of protein in milk/% of transcript in the mammary tissue), showed that α s1 -and β-casein transcripts are translated ca. 3-to 4-fold more efficiently than α s2 -and κ-casein transcripts. This seems to be the rule in the three ruminant species studied. More or less optimal contexts for initiation of translation (Kozak recognition sequence of the start codon) as well as 3' untranslated region (UTR) sequences and length might explain, at least in part, our results. These preliminary results which have now to be confirmed with a larger number of individuals to strengthen our findings and conclusions, provides, however, a rational explanation to the unbalanced casein distribution (approximate proportions 4:1:4:1 for α s1 :α s2 :β:κ, respectively) reported for ruminant milks. The possible effects of specific secondary structures in the 5' and 3' UTRs of casein messengers still have to be considered. milk / casein / mRNA / quantification / untranslated regions (UTRs) / RP-HPLC / real time quantitative PCR / goat / cow / ewe

Gene expression of six major milk proteins in primary bovine mammary epithelial cells isolated from milk during the first twenty weeks of lactation

“…Bevilacqua et al (2006) measured equal proportions of casein gene transcripts which is roughly comparable to our findings. However, the four casein mRNAs were not translated with the same efficiency.…”

“…In lactating dairy ewe mammary gland expression levels of the caseins (CSN1S1, CSN1S2, CSN2, CSN3) did not change, but during involution and late pregnancy the expression levels lowered (Colitti and Pulina, 2010). Bevilacqua et al (2006) investigated the expression levels and the translation efficiency of the four caseins in goat, ewe, and cow. In cows, transcripts seemed to be at the same level of abundance.…”

Effect of β-lactoglobulin and κ-casein genes polymorphism on milk composition in indigenous Zel sheep

“…Casein genes are organised as a tightly linked cluster on ovine chromosome 4 in a 250 kb DNA segment (Bevilacqua et al 2006). The performance of technological processes of cheese production depends on the structure of CSN3 protein (Zaton 1999).…”

Expression profile of caseins, estrogen and prolactin receptors in mammary glands of dairy ewes

“…Caseins comprise a group of four proteins (αs1, αs2, β, and κ) resulting from the expression of four structural genes (CSN1S1, CSN1S2, CSN2 and CSN3, respectively) (Bevilacqua et al, 2006). These proteins represent on average 82% of sheep milk Total Nitrogen (N x 6.38; Pulina and Nudda, 2004), but there is a large variability from one species to another (Miranda et al, 2004).…”