Biology subjects

Lipper, C. H.

Publications and source records attributed to Lipper, C. H..

3 recordsLinked to original sources

bioRxiv2023

DNA binding analysis of rare variants in homeodomains reveals novel homeodomain specificity-determining residues

Homeodomains (HDs) are the second largest class of DNA binding domains (DBDs) among eukaryotic sequence-specific transcription factors (TFs) and play important roles in regulating development, body patterning, and cellular differentiation. Here, we analyzed 92 human HD mutants, including disease-associated variants and variants of unknown significance (VUSs), for their effects on DNA binding activity. Many of the variants altered DNA binding affinity and/or specificity. Biochemical analysis and structural modeling identified 14 novel specificity-determining positions, 5 of which do not contact DNA. The same missense substitution at analogous positions within different HDs often exhibited different effects on DNA binding. Variant effect prediction tools perform moderately well in distinguishing variants with altered binding affinity, but poorly in identifying those with altered specificity. Our results highlight the need for biochemical assays of TF coding variants and prioritize dozens of variants for further investigations into their pathogenicity and development of clinical diagnostics and precision therapies.

genetics↗

bioRxiv2022

Structural Basis for Selective Proteolysis of ADAM10 Substrates at Membrane-Proximal Sites

The endopeptidase ADAM10 is a critical catalyst for regulated proteolysis of key drivers of mammalian development and physiology, and for non-amyloidogenic cleavage of the Alzheimers precursor protein as the primary -secretase. ADAM10 function in vivo requires formation of a complex with a C8-tetraspanin protein, with different ADAM10-C8-tetraspanin complexes having distinct substrate selectivity, yet the basis for such selectivity remains elusive. We present here a cryo-EM structure of a vFab-ADAM10-Tspan15 complex, which shows that Tspan15 binding relieves ADAM10 autoinhibition and positions the enzyme active site about 20 [A] from the plasma membrane for membrane-proximal substrate cleavage. Cell-based assays of N-cadherin shedding establish that the positioning of the active site by the interface between the ADAM10 catalytic domain and the bound tetraspanin influences selection of the preferred cleavage site. Together, these studies reveal the molecular mechanism underlying selective ADAM10 proteolysis at membrane-proximal sites and offer a roadmap for its modulation in disease.

biochemistry↗

bioRxiv2022

Tethered agonist activated ADGRF1 structure reveals molecular preference for Gq signaling

Adhesion G-Protein Coupled Receptors (aGPCRs) have evolved an activation mechanism to translate extracellular force into liberation of a tethered agonist (TA) to modulate cell signalling. We report here that ADGRF1 is the first class B GPCR shown to signal through all major G-protein classes and identify the structural basis for its Gq preference by cryo-EM. Our structure shows that Gq over Gs preference in ADGRF1 derives from tighter packing at the conserved F569 of the TA, altering contacts between TM helix I and VII, with a concurrent rearrangement of TM helices VII and VIII at the site of G recruitment. Gs signalling is also more sensitive to mutation of TA or binding site residues than Gq. Our work advances the understanding of aGPCR TA activation in molecular detail, identifying structural features that potentially explain preferential signal modulation.

biochemistry↗