αS1-casein, which is essential for efficient ER-to-Golgi casein transport, is also present in a tightly membrane-associated form

Annabelle Le Parc,Joëlle Leonil,Eric Chanat

doi:10.1186/1471-2121-11-65

Abstract

BackgroundCaseins, the main milk proteins, aggregate in the secretory pathway of mammary epithelial cells into large supramolecular structures, casein micelles. The role of individual caseins in this process and the mesostructure of the casein micelle are poorly known.ResultsIn this study, we investigate primary steps of casein micelle formation in rough endoplasmic reticulum-derived vesicles prepared from rat or goat mammary tissues. The majority of both αS1- and β-casein which are cysteine-containing casein was dimeric in the endoplasmic reticulum. Saponin permeabilisation of microsomal membranes in physico-chemical conditions believed to conserve casein interactions demonstrated that rat immature β-casein is weakly aggregated in the endoplasmic reticulum. In striking contrast, a large proportion of immature αS1-casein was recovered in permeabilised microsomes when incubated in conservative conditions. Furthermore, a substantial amount of αS1-casein remained associated with microsomal or post-ER membranes after saponin permeabilisation in non-conservative conditions or carbonate extraction at pH11, all in the presence of DTT. Finally, we show that protein dimerisation via disulfide bond is involved in the interaction of αS1-casein with membranes.ConclusionsThese experiments reveal for the first time the existence of a membrane-associated form of αS1-casein in the endoplasmic reticulum and in more distal compartments of the secretory pathway of mammary epithelial cells. Our data suggest that αS1-casein, which is required for efficient export of the other caseins from the endoplasmic reticulum, plays a key role in early steps of casein micelle biogenesis and casein transport in the secretory pathway.

Highlights

Caseins, the main milk proteins, aggregate in the secretory pathway of mammary epithelial cells into large supramolecular structures, casein micelles
Casein micelles vary in size, compactness, protein and mineral compositions, their structure as a whole is believed to be similar across species, implying that very general features are involved in their biogenesis
It follows that the small substructures observed within casein micelles at the electron microscopy level or detected by Small Angle X-ray Scattering as a characteristic point of inflection in SAXS profiles might well be calcium phosphate nanoclusters rather than submicelles [7]

Summary

Introduction

The main milk proteins, aggregate in the secretory pathway of mammary epithelial cells into large supramolecular structures, casein micelles. First proposed by Holt (for review see [2]) and extended by Horne [6], is the tangled web model In this model, caseins self-associate, mainly through hydrophobic and electrostatic interactions, to form a homogeneous network of casein polymers stabilized through interaction with nanoclusters of calcium phosphate. Caseins self-associate, mainly through hydrophobic and electrostatic interactions, to form a homogeneous network of casein polymers stabilized through interaction with nanoclusters of calcium phosphate It follows that the small substructures observed within casein micelles at the electron microscopy level or detected by Small Angle X-ray Scattering as a characteristic point of inflection in SAXS profiles might well be calcium phosphate nanoclusters rather than submicelles [7]. In both models, -casein, which is highly glycosylated, preferentially localizes at the periphery of the micelle and forms a layer at the proteinwater interface, stabilizing the structure and preventing it from aggregating

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