Craig A. Harrison

Antagonists of activin signaling: mechanisms and potential biological applications

Activins are members of the transforming growth factor-beta (TGF-beta) superfamily that control many physiological processes such as cell proliferation and differentiation, immune responses, wound repair and various endocrine activities. Activins elicit these diverse biological responses by signaling via type I and type II receptor serine kinases. Recent studies have revealed details of the roles of inhibin, betaglycan, follistatin and its related protein follistatin-related gene (FLRG), Cripto and BAMBI in antagonizing activin action, and exogenous antagonists against the activin type I (SB-431542 and SB-505124) and type II (activin-M108A) receptors have been developed. Understanding how activin signaling is controlled extracellularly is the first step in providing treatment for wound healing and for disorders such as cachexia and cancer, which result from a deregulated activin pathway.

Cripto forms a complex with activin and type II activin receptors and can block activin signaling

Activin, nodal, Vg1, and growth and differentiation factor 1 are members of the transforming growth factor β superfamily and signal via the activin type II (ActRII/IIB) and type I (ALK4) serine/threonine kinase receptors. Unlike activins, however, signaling by nodal, Vg1, and growth and differentiation factor 1 requires a coreceptor from the epidermal growth factor-Cripto-FRL1-Cryptic protein family such as Cripto. Cripto has important roles during development and oncogenesis and binds nodal or related ligands and ALK4 to facilitate assembly of type I and type II receptor signaling complexes. Because Cripto mediates signaling via activin receptors and binds directly to ALK4, we tested whether transfection with Cripto would affect the ability of activin to signal and/or interact with its receptors. Here we show that Cripto can form a complex with activin and ActRII/IIB. We were unable to detect activin binding to Cripto in the absence of ActRII/IIB, indicating that unlike nodal, activin requires type II receptors to bind Cripto. If cotransfected with ActRII/IIB and ALK4, Cripto inhibited crosslinking of activin to ALK4 and the association of ALK4 with ActRII/IIB. In addition, Cripto blocked activin signaling when transfected into either HepG2 cells or 293T cells. We have also shown that under conditions in which Cripto facilitates nodal signaling, it antagonizes activin. Inhibition of activin signaling provides an additional example of a Cripto effect on the regulation of signaling by transforming growth factor-β superfamily members. Because activin is a potent inhibitor of cell growth in multiple cell types, these results provide a mechanism that may partially explain the oncogenic action of Cripto.

An activin mutant with disrupted ALK4 binding blocks signaling via type II receptors

Activins control many physiologic and pathophysiologic processes in multiple tissues and, like other TGF-β superfamily members, signal via type II (ActRII/IIB) and type I (ALK4) receptor serine kinases. ActRII/IIB are promiscuous receptors known to bind at least a dozen TGF-β superfamily ligands including activins, myostatin, several BMPs, and nodal. Here we utilize a new screening procedure to rapidly identify activin-A mutants with loss of signaling activity. Our goal was to identify activin-A mutants able to bind ActRII but unable to bind ALK4 and which would be, therefore, candidate type II activin receptor antagonists. Using the structure of BMP-2 bound to its type I receptor (ALK3) as a guide, we introduced mutations in the context of the inhibin βA cDNA and assessed the signaling activity of the resulting mutant proteins. We identified several mutants in the finger (M91E, I105E, M108A) and wrist (activin A/activin C chimera, S60P, I63P) regions of activin-A with reduced signaling activity. Of these the M108A mutant displayed the lowest signaling activity while retaining wild-type-like affinity for ActRII. Unlike wild-type activin-A, the M108A mutant was unable to form a cross-linked complex with ALK4 in the presence of ActRII indicating that its ability to bind ALK4 was disrupted. This data suggested that the M108A mutant might be capable of modulating signaling of activin and related ligands. Indeed, the M108A mutant antagonized activin-A and myostatin, but not TGF-β, signaling in 293T cells, indicating it may be generally capable of blocking ligands that signal via ActRII/IIB.

Activins and Inhibins and Their Signaling

Abstract: Activins and inhibins, which were discovered by virtue of their abilities to stimulate or inhibit, respectively, the secretion of FSH, are members of the transforming growth factor‐β (TGFβ) superfamily and exert a broad range of effects on the diffentiation, proliferation and functions of numerous cell types. Activins interact with two structurally related classes of serine/threonine kinase receptors (type I and type II). Inhibin antagonizes activin by binding to the proteoglycan, betaglycan, and forming a stable complex with and, thereby, sequestering type II activin receptors while excluding type I receptors. If betaglycan is present, inhibin can also antagonize those bone morphogenic proteins (BMPs) whose signaling is dependent upon access to type II activin receptors. Recent insights regarding the structures of ligands, receptors and their signaling complexes are providing the basis for the development of therapeutics capable of modulating fertility and numerous pathophysiologic processes.

Modulation of activin and BMP signaling.

Activins and bone morphogenetic proteins (BMPs) elicit diverse biological responses by signaling through two pairs of structurally related types I and II receptors. Here, we summarize recent advances in understanding the mode of action of activins and BMPs, focusing on our elucidation of the crystal structure of BMP-7 in complex with the extracellular domain (ECD) of the activin type II receptor and our identification of a binding site for activin on the type I receptor ALK4. As a consequence of the broad range of activities of activins and BMPs, it is perhaps not surprising that additional mechanisms are continually being discovered through which a cells responsiveness to these ligands is modulated. In this review, we describe novel ways in which the two extracellular cofactors, betaglycan and Cripto, regulate BMP and/or activin signal transduction.

Activins and inhibins: Physiological roles, signaling mechanisms and regulation

Activins and inhibins belong to the transforming growth factor β (TGFβ) superfamily, which also includes the TGF-β (Massague 1998), bone morphogenetic protein (BMP) (Wozney et al. 1988), growth and differentiation (GDF) and nodal-related families (Schier et al. 2000). In human there are now known to be 42 members of the TGF-β superfamily (reviewed in Shi et al. 2003). This review summarizes the physiological roles of activins and inhibins, focusing on activin actions in the central nervous system (CNS). In addition, we outline the molecular basis for activin signal transduction and regulation, emphasizing recent advances regarding the structural basis for ligand/receptor interactions and the roles of betaglycan and Cripto in attenuating activin signaling.