Leveraging Therapeutic Proteins and Peptides from Lumbricus Earthworms: Targeting SOCS2 E3 Ligase for Cardiovascular Therapy through Molecular Dynamics Simulations.

Suppressor of cytokine signaling 2 (SOCS2), an E3 ubiquitin ligase, regulates the JAK/STAT signaling pathway, essential for cytokine signaling and immune responses. Its dysregulation contributes to cardiovascular diseases (CVDs) by promoting abnormal cell growth, inflammation, and resistance to cell death. This study aimed to elucidate the molecular mechanisms underlying the interactions between Lumbricus-derived proteins and peptides and SOCS2, with a focus on identifying potential therapeutic candidates for CVDs. Utilizing a multifaceted approach, advanced computational methodologies, including 3D structure modeling, protein-protein docking, 100 ns molecular dynamics (MD) simulations, and MM/PBSA calculations, were employed to assess the binding affinities and functional implications of Lumbricus-derived proteins on SOCS2 activity. The findings revealed that certain proteins, such as Lumbricin, Chemoattractive glycoprotein ES20, and Lumbrokinase-7T1, exhibited similar activities to standard antagonists in modulating SOCS2 activity. Furthermore, MM/PBSA calculations were employed to assess the binding free energies of these proteins with SOCS2. Specifically, Lumbricin exhibited an average ΔGbinding of -59.25 kcal/mol, Chemoattractive glycoprotein ES20 showed -55.02 kcal/mol, and Lumbrokinase-7T1 displayed -69.28 kcal/mol. These values suggest strong binding affinities between these proteins and SOCS2, reinforcing their potential therapeutic efficacy in cardiovascular diseases. Further in vitro and animal studies are recommended to validate these findings and explore broader applications of Lumbricus-derived proteins.

Molecular insights into degron recognition by CRL5ASB7 ubiquitin ligase.

The ankyrin (ANK) SOCS box (ASB) family, encompassing ASB1-18, is the largest group of substrate receptors of cullin 5 Ring E3 ubiquitin ligase. Nonetheless, the mechanism of substrate recognition by ASB family proteins has remained largely elusive. Here we present the crystal structure of ASB7-Elongin B-Elongin C ternary complex bound to a conserved helical degron. ASB7 employs its ANK3-6 to form an extended groove, effectively interacting with the internal α-helix-degron through a network of side-chain-mediated electrostatic and hydrophobic interactions. Our structural findings, combined with biochemical and cellular analyses, identify the key residues of the degron motif and ASB7 required for their recognition. This will facilitate the identification of additional physiological substrates of ASB7 by providing a defined degron motif for screening. Furthermore, the structural insights provide a basis for the rational design of compounds that can specifically target ASB7 by disrupting its interaction with its cognate degron.

Genome-wide identification and characterization of large yellow croaker (Larimichthys crocea) suppressors of cytokine signaling (SOCS) in immune response to Pseudomonas plecoglossicida infection and acute hypoxia stress.

The suppressor of cytokine signaling (SOCS) gene family is a group of genes involved in the negative regulation of cytokine signal transduction. The members of this family play a crucial role in regulating immune and inflammatory processes. However, comprehensive investigations of these genes have not yet been conducted in the economically significant fish large yellow croaker (Larimichthys crocea). In this study, a total of 13 SOCS genes (LcSOCS1a, LcSOCS1b, LcSOCS2, LcSOCS3a, LcSOCS3b, LcSOCS4, LcSOCS5a, LcSOCS5b, LcSOCS6, LcSOCS7a, LcSOCS7b, LcCISHa and LcCISHb) were identified and analyzed in L. crocea. The phylogenetic tree revealed a high conservation of SOCS genes in evolution, and the gene structure and motif analysis indicated a high similarity in the structure of LcSOCSs in the same subfamily. In addition, the expression patterns of LcSOCSs showed that LcSOCS1b was significantly down-regulated in all time under acute hypoxia stress, but it was markedly up-regulated throughout the entire process after P. plecoglossicida infection, revealing its different immune effects to two stresses. Besides, LcSOCS2a, LcSOCS6 and LcSOCS7a only participated in acute hypoxic stress, while LcSOCS5a was more sensitive to P. plecoglossicida infection. In summary, these results indicated that SOCS genes were involved in stress responses to both biological and non-biological stimuli, setting the foundation for deeper study on the functions of SOCS genes.

Subtle Structural Differences Affect the Inhibitory Potency of RGD-Containing Cyclic Peptide Inhibitors Targeting SPSB Proteins.

The SPRY domain-containing SOCS box proteins SPSB1, SPSB2, and SPSB4 utilize their SPRY/B30.2 domain to interact with a short region in the N-terminus of inducible nitric oxide synthase (iNOS), and recruit an E3 ubiquitin ligase complex to polyubiquitinate iNOS, resulting in the proteasomal degradation of iNOS. Inhibitors that can disrupt the endogenous SPSB-iNOS interactions could be used to augment cellular NO production, and may have antimicrobial and anticancer activities. We previously reported the rational design of a cyclic peptide inhibitor, cR8, cyclo(RGDINNNV), which bound to SPSB2 with moderate affinity. We, therefore, sought to develop SPSB inhibitors with higher affinity. Here, we show that cyclic peptides cR7, cyclo(RGDINNN), and cR9, cyclo(RGDINNNVE), have ~6.5-fold and ~2-fold, respectively, higher SPSB2-bindng affinities than cR8. We determined high-resolution crystal structures of the SPSB2-cR7 and SPSB2-cR9 complexes, which enabled a good understanding of the structure-activity relationships for these cyclic peptide inhibitors. Moreover, we show that these cyclic peptides displace full-length iNOS from SPSB2, SPSB1, and SPSB4, and that their inhibitory potencies correlate well with their SPSB2-binding affinities. The strongest inhibition was observed for cR7 against all three iNOS-binding SPSB proteins.

Ad-hoc modifications of cyclic mimetics of SOCS1 protein: Structural and functional insights.

Suppressors of cytokine signaling 1 (SOCS1) protein, a negative regulator of the Janus kinase (JAK)-signal transducer and activator of transcription (STAT) pathway, possesses a small kinase inhibitory region (KIR) involved in the inhibition of JAK kinases. Several studies showed that mimetics of KIR-SOCS1 can be potent therapeutics in several disorders (e.g., neurological, autoimmune or cardiovascular diseases). In this work, starting from a recently identified cyclic peptidomimetic of KIR-SOCS1, icPS5(Nal1), to optimize the peptide structure and improve its biological activity, we designed novel derivatives, containing crucial amino acids substitutions and/or modifications affecting the ring size. By combining microscale thermophoresis (MST), Circular Dichroism (CD), Nuclear Magnetic Resonance (NMR) and computational studies, we showed that the cycle size plays a key role in the interaction with JAK2 and the substitution of native residues with un-natural building blocks is a valid tool to maintain low-micromolar affinity toward JAK2, greatly increasing their serum stability. These findings contribute to increase the structural knowledge required for the recognition of SOCS1/JAK2 and to progress towards their conversion into more drug-like compounds.

Optimization of Phosphotyrosine Peptides that Target the SH2 Domain of SOCS1 and Block Substrate Ubiquitination.

Suppressor of cytokine signaling 1 (SOCS1) has emerged as a potential therapeutic target in inflammatory and viral diseases. SOCS1 operates via its kinase inhibitory region, Src homology 2 (SH2) domain, and SOCS box to negatively regulate the Janus kinase/signal transducers and activators of transcription signaling pathway. In this study, we utilized native phosphotyrosine peptide substrates as a starting point to iteratively explore the requirement of each amino acid position to target the SH2 domain of SOCS1. We show that Met, Thr, Thr, Val, and Asp in the respective -1, +1, +2, +3, and +5 positions within the peptide substrate are favored for binding to the SOCS1-SH2 domain and identifying several phosphotyrosine peptides that have potent SOCS1 binding affinity with IC50 values ranging from 20 to 70 nM and greater than 100-fold selectivity against the closely related SOCS family proteins, CIS, SOCS2, and SOCS3. The optimized phosphotyrosine peptide was shown to stabilize SOCS1 in a thermal shift assay using cell lysates and inhibited SOCS1-mediated ubiquitination of a target substrate in a biochemical assay. Collectively, these data provide the framework to develop cell-permeable peptidomimetics that further investigate the potential of the SOCS1-SH2 domain as a therapeutic target in inflammatory and viral diseases.

Discovery of an exosite on the SOCS2-SH2 domain that enhances SH2 binding to phosphorylated ligands.

Suppressor of cytokine signaling (SOCS)2 protein is a key negative regulator of the growth hormone (GH) and Janus kinase (JAK)-Signal Transducers and Activators of Transcription (STAT) signaling cascade. The central SOCS2-Src homology 2 (SH2) domain is characteristic of the SOCS family proteins and is an important module that facilitates recognition of targets bearing phosphorylated tyrosine (pTyr) residues. Here we identify an exosite on the SOCS2-SH2 domain which, when bound to a non-phosphorylated peptide (F3), enhances SH2 affinity for canonical phosphorylated ligands. Solution of the SOCS2/F3 crystal structure reveals F3 as an α-helix which binds on the opposite side of the SH2 domain to the phosphopeptide binding site. F3:exosite binding appears to stabilise the SOCS2-SH2 domain, resulting in slower dissociation of phosphorylated ligands and consequently, enhances binding affinity. This biophysical enhancement of SH2:pTyr binding affinity translates to increase SOCS2 inhibition of GH signaling.

Suppressor of cytokine signalling 6 is a potential regulator of antimicrobial peptides in the Chinese oak silkworm, Antheraea pernyi.

The SOCS/CIS is a family of intracellular proteins distributed widely among living organisms. The members of this family have extensively been studied in mammals and have been shown to regulate various physiological processes. In contrast, the functional roles of SOCS/CIS family proteins are unknown in most invertebrates, including insects. Here, we retrieved a full-length open reading frame (ORF) of SOCS-6 from Chines oak silkworm, Antheraea pernyi (Designated as ApSOCS-6), using the RNA-seq database. The predicted ApSOCS-6 amino acid sequence comprised an N-terminal SH2 domain and a C-terminal SOCS-box domain. It shared the highly conserved structures of the SOCS proteins with other lepidopteran species. ApSOCS-6 mRNA transcript was detected in all the tested tissues of the A. pernyi larvae; however, the highest mRNA levels were found in the larval hemocytes, fat bodies, and integuments. The mRNA transcript levels of ApSOCS-6 were increased in the A. pernyi larval hemocytes and fat bodies after a challenge by the Gram-positive bacteria, M. luteus, Gram-negative bacteria, Escherichia coli, Virus, ApNPV, and Fungus, B. bassiana. After the knockdown of ApSOCS-6, we found a significant increase in bacterial clearance and a decrease in the relative replication of bacteria. To evaluate the possible cause of enhanced antibacterial activity, we measured antimicrobial peptides expression in the fat body of A. pernyi larvae. The production of AMPs was strongly increased in the B. cereus infected larval fat bodies following silencing of ApSOCS-6. Our data indicate that ApSOCS-6 negatively regulates the expression of AMPs in immune tissues in response to the B. cereus challenge.

Ankyrin-repeat and SOCS box-containing protein 9 (ASB9) regulates ovarian granulosa cells function and MAPK signaling.

Ankyrin-repeat and SOCS box-containing proteins (ASB) interact with the elongin B-C adapter via their SOCS box domain and with the cullin and ring box proteins to form E3 ubiquitin ligase complexes within the protein ubiquitination pathway. ASB9 in particular is a differentially expressed gene in ovulatory follicles (OFs) induced by the luteinizing hormone (LH) surge or hCG injection in ovarian granulosa cells (GC) while downregulated in growing dominant follicles. Although ASB9 has been involved in biological processes such as protein modification, the signaling network associated with ASB9 in GC is yet to be fully defined. We previously identified and reported ASB9 interactions and binding partners in GC including PAR1, TAOK1, and TNFAIP6/TSG6. Here, we further investigate ASB9 effects on target binding partners regulation and signaling in GC. CRISPR/Cas9-induced inhibition of ASB9 revealed that ASB9 regulates PAR1, TAOK1, TNFAIP6 as well as genes associated with proliferation and cell cycle progression such as PCNA, CCND2, and CCNE2 while CCNA2 was not affected. Inhibition of ASB9 was also associated with increased GC number and decreased caspase3/7 activity, CASP3 expression, and BAX/BCL2 ratio. Furthermore, ASB9 induction in OF in vivo 24 h post-hCG is concomitant with a significant decrease in phosphorylation levels of MAPK3/1 while pMAPK3/1 levels increased following ASB9 inhibition in GC in vitro. Together, these results provide strong evidence for ASB9 as a regulator of GC activity and function by modulating MAPK signaling likely through specific binding partners such as PAR1, therefore controlling GC proliferation and contributing to GC differentiation into luteal cells.

Structural basis for the regulation of inducible nitric oxide synthase by the SPRY domain-containing SOCS box protein SPSB2, an E3 ubiquitin ligase.

Relatively high concentration of nitric oxide (NO) produced by inducible nitric oxide synthase (iNOS) in response to a variety of stimuli is a source of reactive nitrogen species, an important weapon of host innate immune defense. The SPRY domain-containing SOCS box protein 2 (SPSB2) is an E3 ubiquitin ligase that regulates the lifetime of iNOS. SPSB2 interacts with the N-terminal region of iNOS via a binding site on the SPRY domain of SPSB2, and recruits an E3 ubiquitin ligase complex to polyubiquitinate iNOS, leading to its proteasomal degradation. Although critical residues for the SPSB2-iNOS interaction have been identified, structural basis for the interaction remains to be explicitly determined. In this study, we have determined a crystal structure of the N-terminal region of iNOS in complex with the SPRY domain of SPSB2 at 1.24 Å resolution. We have resolved the roles of some flanking residues, whose contribution to the SPSB2-iNOS interaction was structurally unclear previously. Furthermore, we have evaluated the effects of SPSB2 inhibitors on NO production using transient transfection and cell-penetrating peptide approaches, and found that such inhibitors can elevate NO production in RAW264.7 macrophages. These results thus provide a useful basis for the development of potent SPSB2 inhibitors as well as recruiting ligands for proteolysis targeting chimera (PROTAC) design.

The Mechanism of NEDD8 Activation of CUL5 Ubiquitin E3 Ligases.

Cullin RING E3 ligases (CRLs) ubiquitylate hundreds of important cellular substrates. Here we have assembled and purified the Ankyrin repeat and SOCS Box protein 9 CUL5 RBX2 ligase (ASB9-CRL) in vitro and show how it ubiquitylates one of its substrates, CKB. CRLs occasionally collaborate with RING between RING E3 ligases (RBRLs), and indeed, mass spectrometry analysis showed that CKB is specifically ubiquitylated by the ASB9-CRL-ARIH2-UBE2L3 complex. Addition of other E2s such as UBE2R1 or UBE2D2 contributes to polyubiquitylation but does not alter the sites of CKB ubiquitylation. Hydrogen-deuterium exchange mass spectrometry (HDX-MS) analysis revealed that CUL5 neddylation allosterically exposes its ARIH2 binding site, promoting high-affinity binding, and it also sequesters the NEDD8 E2 (UBE2F) binding site on RBX2. Once bound, ARIH2 helices near the Ariadne domain active site are exposed, presumably relieving its autoinhibition. These results allow us to propose a model of how neddylation activates ASB-CRLs to ubiquitylate their substrates.

Ikaros antagonizes DNA binding by STAT5 in pre-B cells.

The IKZF1 gene, which encodes the Ikaros transcription factor, is frequently deleted or mutated in patients with B-cell precursor acute lymphoblastic leukemias that express oncogenes, like BCR-ABL, which activate the JAK-STAT5 pathway. Ikaros functionally antagonizes the transcriptional programs downstream of IL-7/STAT5 during B cell development, as well as STAT5 activity in leukemic cells. However, the mechanisms by which Ikaros interferes with STAT5 function is unknown. We studied the genomic distribution of Ikaros and STAT5 on chromatin in a murine pre-B cell line, and found that both proteins colocalize on >60% of STAT5 target regions. Strikingly, Ikaros activity leads to widespread loss of STAT5 binding at most of its genomic targets within two hours of Ikaros induction, suggesting a direct mechanism. Ikaros did not alter the level of total or phosphorylated STAT5 proteins, nor did it associate with STAT5. Using sequences from the Cish, Socs2 and Bcl6 genes that Ikaros and STAT5 target, we show that both proteins bind overlapping sequences at GGAA motifs. Our results demonstrate that Ikaros antagonizes STAT5 DNA binding, in part by competing for common target sequences. Our study has implications for understanding the functions of Ikaros and STAT5 in B cell development and transformation.

SOCS Proteins Participate in the Regulation of Innate Immune Response Caused by Viruses.

The host immune system has multiple innate immune receptors that can identify, distinguish and react to viral infections. In innate immune response, the host recognizes pathogen-associated molecular patterns (PAMP) in nucleic acids or viral proteins through pathogen recognition receptors (PRRs), especially toll-like receptors (TLRs) and induces immune cells or infected cells to produce type I Interferons (IFN-I) and pro-inflammatory cytokines, thus when the virus invades the host, innate immunity is the earliest immune mechanism. Besides, cytokine-mediated cell communication is necessary for the proper regulation of immune responses. Therefore, the appropriate activation of innate immunity is necessary for the normal life activities of cells. The suppressor of the cytokine signaling proteins (SOCS) family is one of the main regulators of the innate immune response induced by microbial pathogens. They mainly participate in the negative feedback regulation of cytokine signal transduction through Janus kinase signal transducer and transcriptional activator (JAK/STAT) and other signal pathways. Taken together, this paper reviews the SOCS proteins structures and the function of each domain, as well as the latest knowledge of the role of SOCS proteins in innate immune caused by viral infections and the mechanisms by which SOCS proteins assist viruses to escape host innate immunity. Finally, we discuss potential values of these proteins in future targeted therapies.

BC-Box Motif in SOCS6 Induces Differentiation of Epidermal Stem Cells into GABAnergic Neurons.

The BC-box motif in suppressor of cytokine signaling 6 (SOCS6) promotes the neuronal differentiation of somatic stem cells, including epidermal stem cells. SOCS6 protein belongs to the group of SOCS proteins and inhibits cytokine signaling. Here we showed that epidermal stem cells were induced to differentiate into GABAnergic neurons by the intracellular delivery of a peptide composed of the amino-acid sequences encoded by the BC-box motif in SOCS6 protein. The BC-box motif (SLQYLCRFVI) in SOCS6 corresponded to the binding site of elongin BC. GABAnergic differentiation mediated by the BC-box motif in SOCS6 protein was caused by ubiquitination of JAK2 and inhibition of the JAK2-STAT3 pathway. Furthermore, GABAnergic neuron-like cells generated from epidermal stem cells were transplanted into the brain of a rodent ischemic model. Then, we demonstrated that these transplanted cells were GAD positive and that the cognitive function of the ischemic model rodents with the transplanted cells was improved. This study could contribute to not only elucidating the mechanism of GABAnergic neuronal differentiation but also to neuronal regenerative medicine utilizing GABAnergic neurons.

Structure and dynamics of the ASB9 CUL-RING E3 Ligase.

The Cullin 5 (CUL5) Ring E3 ligase uses adaptors Elongins B and C (ELOB/C) to bind different SOCS-box-containing substrate receptors, determining the substrate specificity of the ligase. The 18-member ankyrin and SOCS box (ASB) family is the largest substrate receptor family. Here we report cryo-EM data for the substrate, creatine kinase (CKB) bound to ASB9-ELOB/C, and for full-length CUL5 bound to the RING protein, RBX2, which binds various E2s. To date, no full structures are available either for a substrate-bound ASB nor for CUL5. Hydrogen-deuterium exchange (HDX-MS) mapped onto a full structural model of the ligase revealed long-range allostery extending from the substrate through CUL5. We propose a revised allosteric mechanism for how CUL-E3 ligases function. ASB9 and CUL5 behave as rigid rods, connected through a hinge provided by ELOB/C transmitting long-range allosteric crosstalk from the substrate through CUL5 to the RBX2 flexible linker.

CgSOCS6 negatively regulates the expression of CgIL17s and CgDefh1 in the pacific oyster Crassostrea gigas.

As a family of negatively feedback regulating factors, the suppressor of cytokine signaling (SOCS) can depress cytokine signal transduction, and eventually modulate growth, development, differentiation, and immune response. In the present study, a SOCS homologue (designated as CgSOCS6) was identified from oyster Crassostrea gigas. The open reading frame of CgSOCS6 cDNA was of 1167 bp encoding a peptide of 388 amino acid residues with a central Src homology 2 (SH2) domain, a conserved C-terminal SOCS box, and a nucleus localization sequence (NLS) in its N-terminus. The deduced amino acid sequence of CgSOCS6 shared 37.9-45.5% similarity with other SOCS6/7 family members. In the unrooted phylogenetic tree, CgSOCS6 was clustered with EsSOCS6 from Chinese mitten crab Eriocheir sinensis and assigned into the SOCS6/7 group. The mRNA transcripts of CgSOCS6 were constitutively distributed in all the tested tissues, with the highest level in hemocytes. After lipopolysaccharide (LPS) stimulation, the mRNA expression of CgSOCS6 in hemocytes was significantly up-regulated to the highest level at 6 h (8.48-fold compared to the control group, p < 0.01), and then kept at a relatively higher level from 12 h to 72 h. CgSOCS6 protein could be translocated into the hemocyte nucleus after LPS stimulation. The mRNA expressions of interleukin 17-4 (CgIL17-4), CgIL17-5, and defensin (CgDefh1) in the hemocytes of CgSOCS6-knockdown oysters increased significantly (2.55-fold, 2.68-fold, 4.68-fold of that in EGFP-RNAi oysters, p < 0.05, p < 0.05, p < 0.001, respectively) after LPS stimulation. These findings suggested that CgSOCS6 was involved in the oyster immune response by regulating the expressions of CgIL17-4, CgIL17-5, and CgDefh1.

SPSB2 inhibits hepatitis C virus replication by targeting NS5A for ubiquitination and degradation.

Hepatitis C virus (HCV) replication involves many viral and host factors. Host factor SPRY domain- and SOCS box-containing protein 2(SPSB2) belongs to SPSB family, and it recruits target proteins by the SPRY domain and forms E3 ubiquitin ligase complexes by the SOCS box. As an adaptor protein, it can regulate the host's response to infection, but little is known about whether SPSB2 plays a role in HCV replication. In the present study, we found that HCV infection significantly upregulated the mRNA and protein levels of SPSB2 in HCVcc-infected cells. Exogenous expression of SPSB2 in hepatoma cells decreased HCV RNA and protein levels which depended on the SOCS box, while knockdown of endogenous SPSB2 increased HCV RNA and protein levels. Additionally, we demonstrated that SPSB2 interacted with HCV structural protein E1 and nonstructural protein protein 5A (NS5A) via the C-terminal portion of the SPSB2 SPRY domain. Furthermore, SPSB2 induced NS5A ubiquitination and mediated NS5A degradation. Collectively, this study discovered host factor SPSB2 significantly inhibits HCV replication by interacting and degrading NS5A.

Crystal structure of the SPRY domain of human SPSB2 in the apo state.

The SPRY domain-containing SOCS box protein 2 (SPSB2) is one of four mammalian SPSB proteins that are characterized by a C-terminal SOCS box and a central SPRY/B30.2 domain. SPSB2 interacts with inducible nitric oxide synthase (iNOS) via the SPRY domain and polyubiquitinates iNOS, resulting in its proteasomal degradation. Inhibitors that can disrupt SPSB2-iNOS interaction and augment NO production may serve as novel anti-infective and anticancer agents. The previously determined murine SPSB2 structure may not reflect the true apo conformation of the iNOS-binding site. Here, the crystal structure of human SPSB2 SPRY domain in the apo state is reported at a resolution of 1.9 Å. Comparison of the apo and ligand-bound structures reveals that the iNOS-binding site is highly preformed and that major conformational changes do not occur upon ligand binding. Moreover, the C-terminal His6 tag of the recombinant protein binds to a shallow pocket adjacent to the iNOS-binding site on a crystallographically related SPSB2 molecule. These findings may help in structure-based and fragment-based SPSB2 inhibitor design in the future.

Hepatocyte growth control by SOCS1 and SOCS3.

The extraordinary capacity of the liver to regenerate following injury is dependent on coordinated and regulated actions of cytokines and growth factors. Whereas hepatocyte growth factor (HGF) and epidermal growth factor (EGF) are direct mitogens to hepatocytes, inflammatory cytokines such as TNFα and IL-6 also play essential roles in the liver regeneration process. These cytokines and growth factors activate different signaling pathways in a sequential manner to elicit hepatocyte proliferation. The kinetics and magnitude of these hepatocyte-activating stimuli are tightly regulated to ensure restoration of a functional liver mass without causing uncontrolled cell proliferation. Hepatocyte proliferation can become deregulated under conditions of chronic inflammation, leading to accumulation of genetic aberrations and eventual neoplastic transformation. Among the control mechanisms that regulate hepatocyte proliferation, negative feedback inhibition by the 'suppressor of cytokine signaling (SOCS)' family proteins SOCS1 and SOCS3 play crucial roles in attenuating cytokine and growth factor signaling. Loss of SOCS1 or SOCS3 in the mouse liver increases the rate of liver regeneration and renders hepatocytes susceptible to neoplastic transformation. The frequent epigenetic repression of the SOCS1 and SOCS3 genes in hepatocellular carcinoma has stimulated research in understanding the growth regulatory mechanisms of SOCS1 and SOCS3 in hepatocytes. Whereas SOCS3 is implicated in regulating JAK-STAT signaling induced by IL-6 and attenuating EGFR signaling, SOCS1 is crucial for the regulation of HGF signaling. These two proteins also module the functions of certain key proteins that control the cell cycle. In this review, we discuss the current understanding of the functions of SOCS1 and SOCS3 in controlling hepatocyte proliferation, and its implications to liver health and disease.

Structural insights into substrate recognition by the SOCS2 E3 ubiquitin ligase.

The suppressor of cytokine signaling 2 (SOCS2) acts as substrate recognition subunit of a Cullin5 E3 ubiquitin ligase complex. SOCS2 binds to phosphotyrosine-modified epitopes as degrons for ubiquitination and proteasomal degradation, yet the molecular basis of substrate recognition has remained elusive. Here, we report co-crystal structures of SOCS2-ElonginB-ElonginC in complex with phosphorylated peptides from substrates growth hormone receptor (GHR-pY595) and erythropoietin receptor (EpoR-pY426) at 1.98 Å and 2.69 Å, respectively. Both peptides bind in an extended conformation recapitulating the canonical SH2 domain-pY pose, but capture different conformations of the EF loop via specific hydrophobic interactions. The flexible BG loop is fully defined in the electron density, and does not contact the substrate degron directly. Cancer-associated SNPs located around the pY pocket weaken substrate-binding affinity in biophysical assays. Our findings reveal insights into substrate recognition and specificity by SOCS2, and provide a blueprint for small molecule ligand design.