👤 George Sopko

LINGO-1 is a membrane protein of the central nervous system (CNS) that suppresses myelination of axons. Preclinical studies have revealed that blockade of LINGO-1 function leads to CNS repair in demyelinating animal models. The anti-LINGO-1 antibody Li81 (opicinumab), which blocks LINGO-1 function and shows robust remyelinating activity in animal models, is currently being investigated in a Phase 2 clinical trial as a potential treatment for individuals with relapsing forms of multiple sclerosis (AFFINITY: clinical trial.gov number NCT03222973). Li81 has the unusual feature that it contains two LINGO-1 binding sites: a classical site utilizing its complementarity-determining regions and a cryptic secondary site involving Li81 light chain framework residues that recruits a second LINGO-1 molecule only after engagement of the primary binding site. Concurrent binding at both sites leads to formation of a 2:2 complex of LINGO-1 with the Li81 antigen-binding fragment, and higher order complexes with intact Li81 antibody. To elucidate the role of the secondary binding site, we designed a series of Li81 variant constructs that eliminate it while retaining the classic site contacts. These Li81 mutants retained the high affinity binding to LINGO-1, but lost the antibody-induced oligodendrocyte progenitor cell (OPC) differentiation activity and myelination activity in OPC- dorsal root ganglion neuron cocultures seen with Li81. The mutations also attenuate antibody-induced internalization of LINGO-1 on cultured cortical neurons, OPCs, and cells over-expressing LINGO-1. Together these studies reveal that engagement at both LINGO-1 binding sites of Li81 is critical for robust functional activity of the antibody. Show less

Atherosclerosis is the major cause of coronary artery disease (CAD), and oxidized LDL (oxLDL) is believed to play a key role in the initiation of the atherosclerotic process. Recent studies show that inflammation and autoimmune reactions are also relevant in atherosclerosis. In this study, we examined the association of antibodies against oxLDL (anti-oxLDL) with the severity of CAD in 558 Women's Ischemia Syndrome Evaluation (WISE) study samples (465 whites; 93 blacks) determined by coronary stenosis (< 20%, 20%-49%, > 50% stenosis). We also examined the relationship of anti-oxLDL with serum lipid levels and nine candidate genes including APOE, APOH, APOA5, LPL, LRP1, HL, CETP, PON1, and OLR1. IgM anti-oxLDL levels were significantly higher in the >20% stenosis group than in the ≥ 20% stenosis group in whites (0.69 ± 0.02 vs. 0.64 ± 0.01, respectively; P = 0.02). IgM anti-oxLDL levels correlated significantly with total cholesterol (r² = 0.01; P = 0.03) and LDL cholesterol (r² = 0.017; P = 0.004) in whites. Multiple regression analysis revealed a suggestive association of LPL/S447X single-nucleotide polymorphism (SNP) with both IgG anti-oxLDL (P = 0.02) and IgM anti-oxLDL (P = 0.07), as well as between IgM anti-oxLDL and the OLR1/3'UTR SNP (P = 0.020). Our data suggest that higher IgM anti-oxLDL levels may provide protection against coronary stenosis and that genetic variation in some candidate genes are determinants of anti-oxLDL levels. Show less

START-dependent transcription in Saccharomyces cerevisiae is regulated by two transcription factors SBF and MBF, whose activity is controlled by the binding of the repressor Whi5. Phosphorylation and removal of Whi5 by the cyclin-dependent kinase (CDK) Cln3-Cdc28 alleviates the Whi5-dependent repression on SBF and MBF, initiating entry into a new cell cycle. This Whi5-SBF/MBF transcriptional circuit is analogous to the regulatory pathway in mammalian cells that features the E2F family of G1 transcription factors and the retinoblastoma tumor suppressor protein (Rb). Here we describe genetic and biochemical evidence for the involvement of another CDK, Pcl-Pho85, in regulating G1 transcription, via phosphorylation and inhibition of Whi5. We show that a strain deleted for both PHO85 and CLN3 has a slow growth phenotype, a G1 delay, and is severely compromised for SBF-dependent reporter gene expression, yet all of these defects are alleviated by deletion of WHI5. Our biochemical and genetic tests suggest Whi5 mediates repression in part through interaction with two histone deacetylases (HDACs), Hos3 and Rpd3. In a manner analogous to cyclin D/CDK4/6, which phosphorylates Rb in mammalian cells disrupting its association with HDACs, phosphorylation by the early G1 CDKs Cln3-Cdc28 and Pcl9-Pho85 inhibits association of Whi5 with the HDACs. Contributions from multiple CDKs may provide the precision and accuracy necessary to activate G1 transcription when both internal and external cues are optimal. Show less