Biology subjects

Howard, J. A.

Publications and source records attributed to Howard, J. A..

5 recordsLinked to original sources

bioRxiv2026

GDF3 is an endogenous antagonist of the ActE-ALK7/ACVR2 pathway in adipocytes

Metabolic dysfunction-associated steatotic liver disease (MASLD) and steatohepatitis (MASH) arise, in part, from excessive free fatty acid flux from adipose tissue to the liver. Activin E (ActE, encoded by INHBE) suppresses adipocyte lipolysis through the type I receptor ALK7 (ACVR1C). Loss-of-function variants in INHBE and ACVR1C reduce waist-to-hip ratio in humans, yet genetic knockouts in mice produce insulin resistance and hepatic steatosis, suggesting discrepancies between human and mouse biology. We identify GDF3, a TGF-{beta} superfamily ligand upregulated in obese adipose tissue, as the principal endogenous antagonist of ActE/ALK7 signaling. Human transcriptomic datasets reveal coordinated dysregulation: hepatic INHBE expression and circulating ActE protein are elevated in obesity, while adipose ACVR1C is downregulated and GDF3 is reciprocally upregulated. Using ALK7-selective reporter assays, we show GDF3 inhibits ActE-driven SMAD2/3 signaling as a competitive antagonist rather than the weak agonist previously proposed. ActE suppressed beta-adrenergic-stimulated lipolysis in mouse and human adipocytes and primary human adipose tissue; GDF3 overexpression abolished this effect. In diet-induced obese mice, inducible Gdf3 deletion reduced adipose lipolysis, resolved hepatic steatosis and fibrosis, and improved insulin sensitivity, benefits abolished by Inhbe knockdown, confirming dependence on ActE signaling. Predicted loss-of-function variants in INHBE show only nominal, WHR-dependent associations with type 2 diabetes risk, potentially confounded by hematological effects on HbA1c. Gdf3 deficiency synergized with the clinical-stage anti-activin receptor antibody Bimagrumab to amplify fat-mass loss and improve glucose homeostasis in multiple MASH models. These findings establish GDF3 as an endogenous antagonist of ActE-ALK7 signaling and nominate GDF3 inhibition as a therapeutic strategy for MASLD/MASH.

physiology↗

bioRxiv2025

The crystal structure of the activin B:Fst288 complex and computational insights into the broad antagonistic activity and specificity of follistatin

Members of the TGF-{beta} family regulate diverse biological processes, and their activity is tightly controlled by extracellular antagonists such as follistatin 288 (Fst288). Fst288 potently inhibits activins A and B, GDF8, and GDF11. Among these, ActB is the least structurally and functionally characterized, which limits a full understanding of Fst288s broad activity and its specificity for this TGF-{beta} subgroup. To address this, we solved the crystal structure of the ActB:Fst288 complex. As in other complexes, two Fst288 molecules surround ActB, occluding both receptor-binding sites. However, ActB engages Fst288 differently: its fingers bind most strongly the ND domain, while its acidic fingertips uniquely contact FSD3 disrupting interactions between the two Fst288 molecules and reducing the cooperativity seen in other complexes. Computational analysis revealed that, although ActB exhibits higher interaction enthalpy, entropic penalty lowers its overall affinity compared to ActA, consistent with experimentally measured Kd values. We therefore propose that broad ligand inhibition by Fst288 arises from variations in binding interactions and the utilization of cooperativity when direct contacts are insufficient for high affinity. In cellular contexts, antagonistic effectiveness is further impacted by interactions between complexes and the extracellular matrix, resulting in comparable in vitro IC50 values. Finally, sequence comparisons with non-binding TGF-{beta}s indicate that Fst288 specificity originates from ionic contacts with acidic ligand fingertips which likely initiate recognition, while interactions at the type I and II interfaces stabilize the complex. This study provides deeper insight into Fst288s regulation of activin signaling and paves the way for designing inhibitors with desired selectivity.

molecular biology↗

bioRxiv2024

Structural Basis of Non-Latent Signaling by the Anti-Mullerian Hormone Procomplex

Most TGF{beta} family ligands exist as procomplexes consisting of a prodomain noncovalently bound to a growth factor (GF); Whereas some prodomains confer latency, the Anti-Mullerian Hormone (AMH) prodomain maintains a remarkably high affinity for the GF yet remains active. Using single particle EM methods, we show the AMH prodomain consists of two subdomains: a vestigial TGF{beta} prodomain-like fold and a novel, helical bundle GF-binding domain, the result of an exon insertion 450 million years ago, that engages both receptor epitopes. When associated with the prodomain, the AMH GF is distorted into a strained, open conformation whose closure upon bivalent binding of AMHR2 displaces the prodomain through a conformational shift mechanism to allow for signaling.

molecular biology↗

bioRxiv2023

Heterogeneity in Neutrophil Extracellular Traps from Healthy Human Subjects

Neutrophil Extracellular Traps (NETs), a key component of early defense against microbial infection, are also associated with tissue injury. NET composition has been reported to vary with some disease states, but the composition and variability of NETs across many healthy subjects provides a critical comparison that has not been well investigated. We evaluated NETs from twelve healthy subjects of varying ages isolated from multiple blood draws over a three and one half-year period to delineate the variability in extracellular DNA, protein, enzymatic activities, and susceptibility to protease inhibitors. We calculated correlations for NET constituents and loss of human bronchial epithelial barrier integrity, measured by transepithelial electrical resistance, after NET exposure. We found that although there was some variability within the same subject over time, the mean numbers of neutrophils, protein, LDH, serine protease activities, and cytokines IL-8, IL-1RA, and G-CSF in isolated NETs were consistent across subjects. Total DNA and double stranded DNA content in NETs were different across donors. NETs had little or no TNF, IL-17A, or GM-CSF. NET DNA concentration correlated with increased NET neutrophil elastase activity and higher NET IL-1RA concentrations. NET serine protease activity varied considerably within the same donor from day-to-day. Mean response to protease inhibitors was significantly different across donors. NET DNA concentration correlated best with reductions in barrier integrity of human bronchial epithelia. Defining NET concentration by DNA content correlates with other NET components and reductions in NET-driven epithelial barrier dysfunction, suggesting DNA is a reasonable surrogate measurement for these complex structures in healthy subjects.

cell biology↗

bioRxiv2023

Activin E is a TGFβ ligand that signals specifically through activin receptor-like kinase 7.

Activins are one of the three distinct subclasses within the greater Transforming Growth Factor {beta} (TGF{beta}) superfamily. First discovered for their critical roles in reproductive biology, activins have since been shown to alter cellular differentiation and proliferation. At present, members of the activin subclass include activin A (ActA), ActB, ActC, ActE, and the more distant members myostatin and GDF11. While the biological roles and signaling mechanisms of most activins class members have been well-studied, the signaling potential of ActE has remained largely unknown. Here, we characterized the signaling capacity of homodimeric ActE. Molecular modeling of the ligand:receptor complexes showed that ActC and ActE shared high similarity in both the type I and type II receptor binding epitopes. ActE signaled specifically through ALK7, utilized the canonical activin type II receptors, ActRIIA and ActRIIB, and was resistant to the extracellular antagonists follistatin and WFIKKN. In mature murine adipocytes, ActE invoked a SMAD2/3 response via ALK7, similar to ActC. Collectively, our results establish ActE as an ALK7 ligand, thereby providing a link between genetic and in vivo studies of ActE as a regulator of adipose tissue. SignificanceActivin E is a homodimeric member of the TGF{beta} family belonging to the activin subclass. Currently, the signaling capacity of ActE is unknown due to a lack of reliable reagents to study the protein. Here, we demonstrate that ActE acts as a canonical TGF{beta} ligand that signals through SMAD2/3 in an ALK7-dependent manner, similar to ActC. ActE also utilizes the activin type II receptors, ActRIIA and ActRIIB, to signal and is unable to be antagonized by FS288 and WFIKKN2. This study shows that ActE is a signaling ligand and provides a connection between genetic and in vivo studies that links ActE to adiposity.

biochemistry↗