-The purpose of this review is to give an overview of our current knowledge on the polymorphisms occurring in genes coding for milk proteins and responsible for quantitative variability in their expression, thus influencing the protein composition of livestock ruminant milk. The overall genomic organisation of the 6 main ruminant milk protein genes: a-lactalbumin, b-lactoglobulin and the four caseins (a s1 , a s2 , b and k), their chromosomal location and their expression pattern are first summarised before presenting general mechanisms controlling gene expression both at the transcriptional and the post-transcriptional levels. Polymorphisms found in cis-regulatory elements, mainly within the 5'-flanking region of the genes encoding b-lactoglobulin and a s1 -and a s2 -caseins, have been found, in cattle, to influence their transcription rate. In addition, polymorphisms found in the transcription unit, within intron as well as exon sequences, have been shown to be responsible for defects in the processing of primary transcripts and/or the export of messenger RNA to the cytoplasm. Mutations responsible for the occurrence of premature stop codons in a s1 -and b-casein mRNAs have been shown to be associated both with a decrease in the level of the relevant transcripts and the existence of multiple forms of messengers due to alternative splicing (exon skipping, usage of cryptic splice sites). Such a situation, well-exemplified by the gene encoding a s1 -casein in the goat, may have dramatic biological consequences (secretion pathway, casein micelle structure, fat content, etc.) by modifying the message and accordingly the primary structure of the protein as well as its expression. Since some of these polymorphisms dramatically affect technological properties of milk, including cheese yields and organoleptic characteristics, methods mainly based on the PCR technique have been designed and applied in selection and breeding programmes to improve milk protein quality.gene expression / milk protein / genetic polymorphism / ruminants Reprod. Nutr. Dev. 42 (2002) 433-459 433
SummaryMaintenance of tissue boundaries is crucial for control of metastasis. We describe a new signalling pathway in which epithelial cell disruption can be minimised and thereby restricts epithelial-mesenchymal transgressions. This involves the release of insulin-like growth factor (IGF)-binding protein 5 (IGFBP5) from apoptotic cells, which increases the adhesion of epithelial cells on mesenchymal but not epithelial extracellular matrix (ECM), and involves the direct interaction of IGFBP5 and a2b1 integrins. IGFBP5 also induced cell adhesion to vitronectin in the absence of aVb3 integrin, the vitronectin receptor, again through an a2b1-integrin-dependent action, suggesting that IGFBP5 can induce spreading on matrices, even in the absence of the integrins normally used in this process. Using IGFBP5 mutants we demonstrate that the effect is IGF-independent but requires the heparin-binding domain in the C-terminus of IGFBP5. A truncated mutant containing only the C-terminal of IGFBP5 also induced adhesion. Adhesion induced by IGFBP5 was dependent on Cdc42 and resulted in activation of integrin-linked kinase (ILK) and Akt. Consistent with these changes, IGFBP5 facilitated prolonged cell survival in nutrient-poor conditions and decreased phosphorylation of the stress-activated kinase p38 MAPK (MAPK14). Whereas IGFBP5 enhanced adhesion, it inhibited cell migration, although this was not evident using the truncated Cterminal mutant, suggesting that effects of IGFBP5 on adhesion and migration involve different mechanisms. We anticipate that these responses to IGFBP5 would reduce the metastatic potential of cells.
Fibrosis involves activation of fibroblasts, increased production of collagen and fibronectin and transdifferentiation into contractile myofibroblasts. The process resembles aspects of wound-healing but remains unresolved and can be life-threatening when manifest in the kidneys, lungs and liver, in particular. The causes are largely unknown, but recent suggestions that repetitive micro-injury results in the eventual failure of epithelial cell repair due to replicative senescence are gaining favour. This is consistent with the onset of fibrotic diseases in middle age. Because epithelial injury often involves blood loss, inflammatory responses associated with the fibrotic response have been considered as therapeutic targets. However, this has proved largely unsuccessful and focus is now switching to earlier events in the process. These include EMT (epithelial-mesenchymal transition) and fibroblast activation in the absence of inflammation. TGFbeta1 (transforming growth factor-beta1) induces both EMT and fibroblast activation and is considered to be a major pro-fibrotic factor. Recently, IGFBP-5 [IGF (insulin-like growth factor)-binding protein-5] has also been shown to induce similar effects on TGFbeta1, and is strongly implicated in the process of senescence. It also stimulates migration of peripheral blood mononuclear cells, implicating it in the inflammatory response. In this paper, we examine the evidence for a role of IGFBP-5 in fibrosis and highlight its structural relationship with other matrix proteins and growth factors also implicated in tissue remodelling.
Insulin-like growth factor binding proteins (IGFBPs) -3 and -5 are known to interact with various components of the extracellular matrix (ECM; e.g. heparin and heparan sulphate) and this interaction is believed to affect the affinity of both IGFBP species for their cognate ligands -IGF-I and -II. There is little detail on the nature of the molecular complex formed between ECM components, IGFBPs and IGFs although the glycosaminoglycan (GAG) heparin has been reported to reduce the affinity of IGFBP-5 for IGF-I. In order to investigate this phenomenon further, we have undertaken an extensive surface plasmon resonance based biosensor study to report the affinity of IGFBP-3 and -5 for binding heparin (22 and 7 nM respectively). We have also shown that pre-complexation of IGFBP with IGF-I and -II inhibits the subsequent association of IGFBP with heparin and conversely that heparin complexation of IGFBP-3 and -5 inhibits IGFBP binding to biosensor surfaces containing immobilised IGF-I. In addition we have used both IGF-I and heparin coated biosensor surfaces in an attempt to build ternary IGF-IGFBP-heparin complexes in order to gain some insight into the nature of inhibition by heparin of IGFI-IGFBP complex formation. Our data lead us to conclude that the inhibition by heparin is partly competitive in nature, and that ternary complexes of IGF-IGFBP-heparin are either unable to form, or only form unstable transient complexes. The potential biological significance of our data is highlighted by the demonstration that IGF-I and IGF-II can displace endogenous IGFBP-5 from monolayer cultures of the mouse mammary epithelial cell line HC11.
We have reported previously that mutation of two conserved nonbasic amino acids (G203 and Q209) within the highly basic 201-218 region in the C-terminal domain of IGF-binding protein-5 (IGFBP-5) decreases binding to IGFs. This study reveals that cumulative mutagenesis of the 10 basic residues in this region, to create the C-Term series of mutants, ultimately results in a 15-fold decrease in the affinity for IGF-I and a major loss in heparin binding. We examined the ability of mutants to inhibit IGF-mediated survival of MCF-7 cells and were able to demonstrate that this depended not only upon the affinity for IGF-I, but also the kinetics of this interaction, because IGFBP-5 mutants with similar affinity constants (K(D)) values, but with different association (Ka) and dissociation (Kd) rate values, had markedly different inhibitory properties. In contrast, the affinity for IGF-I provided no predictive value in terms of the ability of these mutants to enhance IGF action when bound to the substratum. Instead, these C-Term mutants appeared to enhance the actions of IGF-I by a combination of increased dissociation of IGF-IGFBP complexes from the substratum, together with dissociation of IGF-I from IGFBP-5 bound to the substratum. These effects of the IGFBPs were dependent upon binding to IGF-I, because a non-IGF binding mutant (N-Term) was unable to inhibit or enhance the actions of IGF-I. These results emphasize the importance of the kinetics of association/dissociation in determining the enhancing or inhibiting effects of IGFBP-5 and demonstrate the ability to generate an IGFBP-5 mutant with exclusively IGF-enhancing activity.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
customersupport@researchsolutions.com
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
This site is protected by reCAPTCHA and the Google Privacy Policy and Terms of Service apply.
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.