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Mortensen, J. S.

Publications and source records attributed to Mortensen, J. S..

3 recordsLinked to original sources

MORPHIS (MORPHological Interpretable Signature) captures heterogeneous treatment- and aging-related responses of single cells

Cell morphology encodes changes in cytoskeletal and organelle organization during disease, treatment, and aging, yet is often assessed qualitatively or through poorly interpretable feature sets extracted from microscopy images. Here we introduce MORPHIS (MORPHological Interpretable Signature), a machine learning framework on explainable, analytically rich features paired with statistical methodologies for robust and interpretable quantification of single-cell morphological signature. MORPHIS extracts compact, interpretable feature signatures that capture both perturbation-specific response magnitude and heterogeneous cellular responses. It accurately distinguishes treatment-specific morphological signatures of eight mechanistically distinct membrane-active and intracellular-targeting compounds in Caco-2 and HeLa cells elucidating conserved and divergent phenotypic responses among compound-classes, as well as ultrastructural nuclear alterations upon aging of C. elegans and quantifies heterogeneous single-cell fractional responses. By remaining cell-type and perturbation agnostic, MORPHIS provides a generalizable framework for quantifying morphological signatures across diverse biological contexts including pharmacological treatment or aging.

biophysics↗

Structural basis for selective inhibition of human GABA transporter GAT3

The astrocytic {gamma}-aminobutyric acid (GABA) transporter, GAT3, is essential for terminating GABAergic signalling in the central nervous system. Selective inhibition of GAT3 offers a potential strategy for elevating extracellular GABA levels for the treatment of neurological disorders including epilepsy. However, few potent and selective GAT3 inhibitors have been developed, and their mechanisms of inhibition remain poorly understood. Here, we present the cryo-electron microscopy structures of full-length, wild-type human GAT3 bound to a selective inhibitor, to substrate GABA, or in substrate-free state. GAT3 bound to the inhibitor or in the substrate-free state exhibits an inward-open conformation. The inhibitor binds within the intracellular permeation pathway, positioned between transmembrane helices 1, 2, 3, 6, 7, and 8. The GABA-bound GAT3 is captured in an inward-occluded state, revealing the ion coordination and substrate recognition network, including a cation-{pi} interaction between GABAs {gamma}-amino group and a phenylalanine residue in transmembrane helix 6. Our data reveal the molecular determinants for the inhibitor selectivity, and the mode of substrate binding and transport inhibition, providing blueprints for the rational design of next-generation selective GAT3 inhibitors.

molecular biology↗

Allosteric factors in the calcium/calmodulin-responsive kinase II hub domain determine selectivity of GHB ligands for CaMKIIa

The Ca2+/CaM-dependent protein kinase II alpha (CaMKII) is a highly important synaptic protein, which comprises a unique holoenzyme structure organized via the central hub domain. Recently, a distinct binding pocket in the CaMKII hub domain was identified for the endogenous neuromodulator {gamma}-hydroxybutyric acid (GHB) and related synthetic analogues. Key interacting residues in CaMKII were revealed, but the pronounced selectivity towards the alpha variant of CaMKII has remained unresolved. Aimed at elucidating the molecular determinants for this selectivity, we here conducted binding studies to CaMKII-HEK whole-cell homogenates using two different in-house-developed GHB-related radioligands, [3H]HOCPCA and [3H]O-5-HDC, in combination with site-directed mutagenesis. Binding to CaMKII with the smaller-type radioligand [3H]HOCPCA validated key involvement of the four known residues (His395, Arg433, Arg453 and Arg469), but also revealed a role for the upper hub flexible loop containing the CaMKII-specific residue Trp403 (Leu in all other CaMKII isozymes). Insertion of the corresponding residues (L467W/C533R) into CaMKII{beta} failed to introduce [3H]HOCPCA binding. However, with the larger-type radioligand, [3H]O-5-HDC, specific binding in CaMKII{beta} (L467W/C533R) was achieved. Thus, of the four native CaMKII isozymes, only CaMKII accommodates GHB ligands. The study identifies the CaMKII flexible pocket loop as a distantly located "allosteric" factor in determining selectivity of GHB analogues for CaMKII. It sheds light on a remarkable interplay of the entire hub cavity for accommodation of ligands, and corroborates GHB analogues as CaMKII-selective.

biochemistry↗