Biology subjects

Bianchet, M. A.

Publications and source records attributed to Bianchet, M. A..

3 recordsLinked to original sources

bioRxiv2025

Engineering substrate selectivity in the human sodium/iodide symporter (NIS)

Iodide (I-) uptake mediated by the Na/I- symporter (NIS) is the first step in the biosynthesis of the thyroid hormones, of which I- is an essential constituent. NIS couples the inward transport of I- against its electrochemical gradient to the inward translocation of Na+ down its electrochemical gradient. NIS also transports oxyanions (XO4-s), such as perrhenate (ReO4-) and the environmental pollutant perchlorate (ClO4-). Furthermore, NIS is the basis for radioiodide ({superscript 1}3{superscript 1}I-) therapy for thyroid cancer (administered after thyroidectomy), the most effective targeted internal radiation cancer therapy available. 131I- selectively targets remnant malignant cells and metastases expressing NIS, causing only minor side effects. There is great interest in expressing NIS exogenously, by gene transfer, in extrathyroidal cancers to render them susceptible to destruction by 131I-. This approach, however, would also harm patients thyroids. Therefore, a strategy is needed for killing non-thyroidal cancer cells exogenously expressing NIS while protecting the thyroid. Addressing this need, we present here an engineered double mutant, L253P/V254F (PF)-NIS, which selectively transports XO4-s but not I-. We used cryo-EM to determine the structure of PF-NIS with ReO4- and Na+ ions bound to it at a 2.58 [A] resolution, and showed that high concentrations of non-radioactive I- protect WT-NIS-expressing cells from radioactive 186ReO4-, whereas PF-NIS-expressing cells are killed. Thus, PF-NIS could potentially be used, together with 186/188ReO4- and non-radioactive I-, to treat non-thyroidal cancers while safeguarding the thyroid. This study establishes a framework for developing therapies using NIS molecules engineered to have selective substrate specificities to extend the clinical use of NIS beyond thyroid cancer.

biophysics↗

bioRxiv2023

HCN channels sense temperature and determine heart rate responses to heat

Heart rate increases with heat, [1-3] constituting a fundamental physiological relationship in vertebrates. Each normal heartbeat is initiated by an action potential generated in a sinoatrial nodal pacemaker cell. Pacemaker cells are enriched with hyperpolarization activated cyclic nucleotide-gated ion channels (HCN) that deliver cell membrane depolarizing inward current that triggers action potentials. HCN channel current increases due to cAMP binding, a mechanism coupling adrenergic tone to physiological fight or flight heart rate acceleration. However, the mechanism(s) for heart rate response to thermal energy is unknown. We used thermodynamical and homology computational modeling, site-directed mutagenesis and mouse models to identify a concise motif on the S4-S5 linker of the cardiac pacemaker HCN4 channels (M407/Y409) that determines HCN4 current (If) and cardiac pacemaker cell responses to heat. This motif is required for heat sensing in cardiac pacemaker cells and in isolated hearts. In contrast, the cyclic nucleotide binding domain is not required for heat induced HCN4 current increases. However, a loss of function M407/Y409 motif mutation prevented normal heat and cAMP responses, suggesting that heat sensing machinery is essential for operating the cAMP allosteric pathway and is central to HCN4 modulation. The M407/Y409 motif is conserved across all HCN family members suggesting that HCN channels participate broadly in coupling heat to changes in cell membrane excitability.

physiology↗

bioRxiv2022

Structural insights into sodium/iodide symporter (NIS) substrate binding and specificity

The sodium/iodide symporter (NIS) is the essential plasma membrane protein that mediates active iodide (I-) transport into the thyroid gland, the first step in the biosynthesis of the thyroid hormones--the master regulators of intermediary metabolism. NIS couples the inward translocation of I- against its electrochemical gradient to the inward transport of Na+ down its electrochemical gradient. For nearly 50 years before its molecular identification, NIS was already the molecule at the center of the single most effective internal radiation cancer therapy ever devised: radioiodide (131I-) treatment for thyroid cancer. Mutations in NIS cause congenital hypothyroidism, which must be treated immediately after birth to prevent stunted growth and cognitive deficiency. To date, the structure of NIS has been unknown. Here, we report three structures of rat NIS, determined by single-particle cryo-electron microscopy (cryo-EM): one with no substrates bound, one with 2 Na+ and 1 I- bound, and one with 1 Na+ and the oxyanion perrhenate bound. Structural analyses, functional characterization, and computational studies reveal the substrate binding sites and residues key for transport activity. Our results yield insights into how NIS selects, couples, and translocates anions--thereby establishing a framework for understanding NIS function--and into how it transports different substrates with different stoichiometries and releases substrates from its substrate-binding cavity into the cytosol.

biophysics↗