Biological functions of the small leucine-rich proteoglycans: from genetics to signal transduction

The small leucine-rich proteoglycan (SLRP) family has significantly expanded in the past decade to now encompass five discrete classes, grouped by common structural and functional properties. Some of these gene products are not classical proteoglycans, whereas others have new and unique features. In addition to being structural proteins, SLRPs constitute a network of signal regulation: being mostly extracellular, they are upstream of multiple signaling cascades. They affect intracellular phosphorylation, a major conduit of information for cellular responses, and modulate distinct pathways, including those driven by bone morphogenetic protein/transforming growth factor beta superfamily members, receptor tyrosine kinases such as ErbB family members and the insulin-like growth factor I receptor, and Toll-like receptors. The wealth of mechanistic insights into the molecular and cellular functions of SLRPs has revealed both the sophistication of this family of regulatory proteins and the challenges that remain in uncovering the totality of their functions. This review is focused on novel biological functions of SLRPs with special emphasis on their protein cores, newly described genetic diseases, and signaling events in which SLRPs play key functions.     

Calcium-binding proteins: intracellular sensors from the calmodulin superfamily.

In all eukaryotic cells, and particularly in neurons, Ca(2+) ions are important second messengers in a variety of cellular signaling pathways. In the retina, Ca(2+) modulation plays a crucial function in the development of the visual system's neuronal connectivity and a regulatory role in the conversion of the light signal received by photoreceptors into an electrical signal transmitted to the brain. Therefore, the study of retinal Ca(2+)-binding proteins, which frequently mediate Ca(2+) signaling, has given rise to the important discovery of two subfamilies of these proteins, neuronal Ca(2+)-binding proteins (NCBPs) and calcium-binding proteins (CaBPs), that display similarities to calmodulin (CaM). These and other Ca(2+)-binding proteins are integral components of cellular events controlled by Ca(2+). Some members of these subfamilies also play a vital role in signal transduction outside of the retina. The expansion of the CaM-like protein family reveals diversification among Ca(2+)-binding proteins that evolved on the basis of the classic molecule, CaM. A large number of NCBP and CaBP subfamily members would benefit from their potentially specialized role in Ca(2+)-dependent cellular processes. Pinpointing the role of these proteins will be a challenging task for further research.     

TULA proteins as signaling regulators

Two members of the UBASH3/STS/TULA family exhibit a unique protein domain structure, which includes a histidine phosphatase domain, and play a key role in regulating cellular signaling. UBASH3A/STS-2/TULA is mostly a lymphoid protein, while UBASH3B/STS-1/TULA-2 is expressed ubiquitously. Dephosphorylation of tyrosine-phosphorylated proteins by TULA-2 and, probably to a lesser extent, by TULA critically contribute to the molecular basis of their regulatory effect. The notable differences between the effects of the two family members on cellular signaling and activation are likely to be linked to the difference between their specific enzymatic activities. However, these differences might also be related to the functions of their domains other than the phosphatase domain and independent of their phosphatase activity. The down-regulation of the Syk/Zap-70-mediated signaling, which to-date appears to be the best-studied regulatory effect of TULA family, is discussed in detail in this publication.     

Regulatory roles and molecular signaling of TNF family members in osteoclasts

The tumor necrosis factor (TNF) family has been one of the most intensively studied families of proteins in the past two decades. The TNF family constitutes 19 members that mediate diverse biological functions in a variety of cellular systems. The TNF family members regulate cellular functions through binding to membrane-bound receptors belonging to the TNF receptor (TNFR) family. Members of the TNFR family lack intrinsic kinase activity and thus they initiate signaling by interacting intracellular signaling molecules such as TNFR associated factor (TRAF), TNFR associated death domain (TRADD) and Fas-associated death domain (FADD). In bone metabolism, it has been shown that numerous TNF family members including receptor activator of nuclear factor kappaB ligand (RANKL), TNF-alpha, Fas ligand (FasL) and TNF-related apoptosis-inducing ligand (TRAIL) play pivotal roles in the differentiation, function, survival and/or apoptosis of osteoclasts, the principal bone-resorbing cells. These TNF family members not only regulate physiological bone remodeling but they are also implicated in the pathogenesis of various bone diseases such as osteoporosis and bone loss in inflammatory conditions. This review will focus on our current understanding of the regulatory roles and molecular signaling of these TNF family members in osteoclasts.     

Argonaute proteins: mediators of RNA silencing

Small regulatory RNAs such as short interfering RNAs (siRNAs), microRNAs (miRNAs), and Piwi interacting RNAs (piRNAs) have been discovered in the past, and it is becoming more and more apparent that these small molecules have key regulatory functions. Small RNAs are found in all higher eukaryotes and play important roles in cellular processes as diverse as development, stress response, or transposon silencing. Soon after the discovery of small regulatory RNAs, members of the Argonaute protein family were identified as their major cellular protein interactors. This review focuses on the various cellular functions of mammalian Argonaute proteins in conjunction with the different small RNA species that are known today.     

Receptor tyrosine kinases.

A major process through which environmental information is transmitted into cells is via activation of protein tyrosine kinases. Receptor tyrosine kinases contain extracellular ligand recognition, single membrane spanning, and cytoplasmic protein tyrosine kinase domains. The cytoplasmic kinase core is flanked by regulatory segments, which in some family members are also inserted into the core kinase domain. Ligand binding initiates receptor signaling from the cell surface. Activated receptors autophosphorylate to remove alternate substrate/inhibitory constraints and to provide loci for assembly of proteins that contain SRC homology regions. Information is transmitted and diffused by tyrosine phosphorylation of the assembled proteins and of cellular substrates that include protein kinases with specificity for serine/threonine residues. Signaling, which is strictly ligand-dependent, is attenuated by down-regulation of receptors and by feed-back inhibitory loops that involve receptor phosphorylation by cellular kinases. The tyrosine kinase receptors are essential for normal growth, development, and reparative processes. Mutations that remove normal regulatory constraints on the approximately 290 amino acid kinase core of these large proteins result in constitutive function and cell transformation.     

Functions of heparan sulfate proteoglycans in cell signaling during development

Heparan sulfate proteoglycans (HSPGs) are cell-surface and extracellular matrix macromolecules that are composed of a core protein decorated with covalently linked glycosaminoglycan (GAG) chains. In vitro studies have demonstrated the roles of these molecules in many cellular functions, and recent in vivo studies have begun to clarify their essential functions in development. In particular, HSPGs play crucial roles in regulating key developmental signaling pathways, such as the Wnt, Hedgehog, transforming growth factor-beta, and fibroblast growth factor pathways. This review highlights recent findings regarding the functions of HSPGs in these signaling pathways during development.     

Kinase insert domain receptor (KDR) extracellular immunoglobulin-like domains 4-7 contain structural features that block receptor dimerization and vascular endothelial growth factor-induced signaling

The vascular endothelial growth factor (VEGF) receptor tyrosine kinase subtype kinase insert domain receptor (KDR) contains seven extracellular Ig-like domains, of which the three most amino-terminal contain the necessary structural features required for VEGF binding. To clarify the functional role of KDR Ig-like domains 4-7, we compared VEGF-induced signaling in human embryonic kidney and porcine aortic endothelial cells expressing native versus mutant receptor proteins in which Ig-like domains 4-7, 4-6, or 7 had been deleted. Western blotting using an anti-receptor antibody indicated equivalent expression levels for each of the recombinant proteins. As expected, VEGF treatment robustly augmented native receptor autophosphorylation. In contrast, receptor autophosphorylation, as well as downstream signaling events, were VEGF-independent for cells expressing mutant receptors. (125)I-VEGF(165) bound with equal or better affinity to mutant versus native receptor, although the number of radioligand binding sites was significantly reduced because a significant percentage of mutant, but not native, receptors were localized to the cell interior. As was the case for native KDR, (125)I-VEGF(165) binding to the mutant receptors was dependent upon cell surface heparan sulfate proteoglycans, and (125)I-VEGF(121) bound with an affinity equal to that of (125)I-VEGF(165) to the native and mutant receptors. It is concluded that KDR Ig-like domains 4-7 contain structural features that inhibit receptor signaling by a mechanism that is independent of neuropilin-1 and heparan sulfate proteoglycans. We speculate that this provides a cellular mechanism for blocking unwanted signaling events in the absence of VEGF.     

Regulatory role of CCN3 in melanoma cell interaction with the extracellular matrix

It is increasingly clear that melanoma cells modify their environment not only through the release of growth factors (GFs) and cytokines that have autocrine or paracrine effects and strongly modulate the immune response, but also by secreting proteins that become structural or transient components of the extracellular matrix (ECM). Melanoma cell secreted proteins play a significant role in cell–ECM interactions, helping tumor cells to invade neighbouring stroma, disseminate and survive in other tissue contexts. CCN3/NOV (nephroblastoma overexpressed) is a matricellular protein that belongs to the CCN family of proteins containing six members in humans. Its structure consists of modules related to functional domains previously identified in major regulatory proteins: insulin-like growth factor-binding protein (IGFBP), von Willebrand factor type C repeats (VWC), thrombospondin type 1 repeats, and secreted regulatory factors containing cysteine knot motifs. Extensive studies have indicated that the biological properties of CCN3 are dependent upon the cellular context, and its role in melanoma seems to recapitulate cell context functions.     

Small leucine rich proteoglycan family regulates multiple signalling pathways in neural development and maintenance

The small leucine-rich repeat proteoglycan (SLRPs) family of proteins currently consists of five classes, based on their structural composition and chromosomal location. As biologically active components of the extracellular matrix (ECM), SLRPs were known to bind to various collagens, having a role in regulating fibril assembly, organization and degradation. More recently, as a function of their diverse proteins cores and glycosaminoglycan side chains, SLRPs have been shown to be able to bind various cell surface receptors, growth factors, cytokines and other ECM components resulting in the ability to influence various cellular functions. Their involvement in several signaling pathways such as Wnt, transforming growth factor-β and epidermal growth factor receptor also highlights their role as matricellular proteins. SLRP family members are expressed during neural development and in adult neural tissues, including ocular tissues. This review focuses on describing SLRP family members involvement in neural development with a brief summary of their role in non-neural ocular tissues and in response to neural injury.     

The roles of G proteins in the activation of TRPC4 and TRPC5 transient receptor potential channels

TRPC4 and TRPC5 channels are important regulators of electrical excitability in both gastrointestinal myocytes and neurons. Much is known regarding the assembly and function of these channels including TRPC1 as a homotetramer or a heteromultimer and the roles that their interacting proteins play in controlling these events. Further, they are one of the best-studied targets of G protein-coupled receptors and growth factors in general and Gαq protein coupled receptor or epidermal growth factor in particular. However, our understanding of the roles of Gαi/o proteins on TRPC4/5 channels is still rudimentary. We discuss potential roles for Gαi/o proteins in channel activation in addition to their known role in cellular signaling.     

Cell cycle and apoptosis regulatory protein-1: a novel regulator of apoptosis in the colonic mucosa during aging

Although the regulatory mechanisms for the age-related rise in proliferation and reduction in apoptosis in the colonic mucosa are yet to be fully delineated, we have demonstrated that these events are associated with increased expression and activation of epithelial growth factor receptor (EGFR)/ErbB-1 and some of its receptor family members (EGFRs), indicating their involvement in these processes. However, the downstream signaling events of EGFR and/or its family members regulating age-related changes in mucosal proliferation and apoptosis remain to be delineated. Cell cycle and apoptosis regulatory protein-1 (CARP-1), a novel growth signaling regulator that we isolated, participates in EGFR-dependent signaling. In the current investigation, we examined the involvement of CARP-1 in colonic mucosal growth-related processes during aging. We report that the age-related reduction in apoptosis in the colonic mucosa is associated with increased expression and tyrosine phosphorylation of not only EGFR but also ErbB-2 and ErbB-3. In contrast, protein and mRNA levels of CARP-1 as well as tyrosine phosphorylation of CARP-1 are decreased. Additionally, we have observed that administration of wortmannin, an inhibitor of phosphatidylinositol 3-kinase activity that accelerates apoptosis in the colonic mucosa of aged rats, causes a marked increase in expression and tyrosine phosphorylation of CARP-1. The age-related decline in CARP-1 expression could partly be attributed to increased methylation of the CARP-1 promoter. Taken together, our data suggest that not only EGFR but also its other members are involved in regulating colonic mucosal growth during aging and that CARP-1 may play a crucial role in transducing EGFRs signals.     

Functional distinctions of protein kinase B/Akt isoforms defined by their influence on cell migration

The three mammalian members of the protein kinase B/Akt (PKB/Akt) family have been implicated in a plethora of cellular signaling processes with key functions in control of cellular metabolism, growth, proliferation and apoptosis. As a major target of phosphatidylinositol (PI) 3-kinase signaling, the PKB/Akt isoforms also have central roles in a variety of human cancers, with effects on tumor initiation, progression and metastasis. It has been shown that isoform-specific functions of PKB/Akt family members can contribute to tumorigenesis on multiple levels. A series of recent studies documents the isoform-specific functions of PKB/Akt family members in regulation of cellular motility and migration by influencing numerous cellular targets involved in organization of the actin cytoskeleton, cellular interaction with the extracellular matrix, expression of motility genes and establishment of cellular polarity. A thorough insight into the isoform-specific roles of PKB/Akt proteins is essential for a full understanding of the complex biological outcomes elicited by PI 3-kinase and PKB/Akt signaling.     

The roles of G proteins in the activation of TRPC4 and TRPC5 transient receptor potential channels

TRPC4 and TRPC5 channels are important regulators of electrical excitability in both gastrointestinal myocytes and neurons. Much is known regarding the assembly and function of these channels including TRPC1 as a homotetramer or a heteromultimer and the roles that their interacting proteins play in controlling these events. Further, they are one of the best-studied targets of G protein-coupled receptors and growth factors in general and Gαq protein coupled receptor or epidermal growth factor in particular. However, our understanding of the roles of Gαi/o proteins on TRPC4/5 channels is still rudimentary. We discuss potential roles for Gαi/o proteins in channel activation in addition to their known role in cellular signaling.     

The "Gab" in signal transduction

Tyrosine phosphorylation plays an important role in controlling cellular growth, differentiation and function. Abnormal regulation of tyrosine phosphorylation can result in human diseases such as cancer. A major challenge of signal transduction research is to determine how the initial activation of protein-tyrosine kinases (PTKs) by extracellular stimuli triggers multiple downstream signaling cascades, which ultimately elicit diverse cellular responses. Recent studies reveal that members of the Gab/Dos subfamily of scaffolding adaptor proteins (hereafter, "Gab proteins") play a crucial role in transmitting key signals that control cell growth, differentiation and function from multiple receptors. Here, we review the structure, mechanism of action and function of these interesting molecules in normal biology and disease.     

Synergistic control of cellular adhesion by transmembrane 9 proteins

The transmembrane 9 (TM9) family of proteins contains numerous members in eukaryotes. Although their function remains essentially unknown in higher eukaryotes, the Dictyostelium discoideum Phg1a TM9 protein was recently reported to be essential for cellular adhesion and phagocytosis. Herein, the function of Phg1a and of a new divergent member of the TM9 family called Phg1b was further investigated in D. discoideum. The phenotypes of PHG1a, PHG1b, and PHG1a/PHG1b double knockout cells revealed that Phg1a and Phg1b proteins play a synergistic but not redundant role in cellular adhesion, phagocytosis, growth, and development. Complementation analysis supports a synergistic regulatory function rather than a receptor role for Phg1a and Phg1b proteins. Together, these results suggest that Phg1 proteins act as regulators of cellular adhesion, possibly by controlling the intracellular transport in the endocytic pathway and the composition of the cell surface.     

胰岛素受体底物家族与Ⅱ型糖尿病

胰岛素受体底物(insulin receptor substrate,IRS)家族是胰岛素／类胰岛素生长因子信号系统上游通路的关键介导者,在维持细胞生长、分裂和代谢中起着重要作用。已有四个成员被鉴定出：IRS-1、IRS-2、IRS-3和IRS-4,其中IRS-1和IRS-2在许多不同的组织细胞中起着特异性作用。IRS介导的胰岛素信号通路与很多其他信号通路存在交叉,它们能干扰胰岛素发挥效应,导致胰岛素抵抗,从而引发糖尿病。     

A spoonful of sugar makes the melanoma go: the role of heparan sulfate proteoglycans in melanoma metastasis

Heparan sulfate proteoglycans (HSPGs) have been shown to regulate signaling in many systems and are of increasing interest in cancer. While these are not the only sugars to drive melanoma metastasis, HSPGs play important roles in driving metastatic signaling cascades in melanoma. The ability of these proteins to modulate ligand-receptor interactions in melanoma has been quite understudied. Recent data from several groups indicate the importance of these ligands in modulating key signaling pathways including Wnt and fibroblast growth factor (FGF) signaling. In this review, we summarize the current knowledge regarding the structure and function of these proteoglycans and their role in melanoma. Understanding how HSPGs modulate signaling in melanoma could lead to new therapeutic approaches via the dampening or heightening of key signaling pathways.