The CaMKII/MLC1 Axis Confers Ca2+-Dependence to Volume-Regulated Anion Channels (VRAC) in Astrocytes

Abstract

Astrocytes, the main glial cells of the central nervous system, play a key role in brain volume control due to their intimate contacts with cerebral blood vessels and the expression of a distinctive equipment of proteins involved in solute/water transport. Among these is MLC1, a protein highly expressed in perivascular astrocytes and whose mutations cause megalencephalic leukoencephalopathy with subcortical cysts (MLC), an incurable leukodystrophy characterized by macrocephaly, chronic brain edema, cysts, myelin vacuolation, and astrocyte swelling. Although, in astrocytes, MLC1 mutations are known to affect the swelling-activated chloride currents (ICl,swell) mediated by the volume-regulated anion channel (VRAC), and the regulatory volume decrease, MLC1’s proper function is still unknown. By combining molecular, biochemical, proteomic, electrophysiological, and imaging techniques, we here show that MLC1 is a Ca2+/Calmodulin-dependent protein kinase II (CaMKII) target protein, whose phosphorylation, occurring in response to intracellular Ca2+ release, potentiates VRAC-mediated ICl,swell. Overall, these findings reveal that MLC1 is a Ca2+-regulated protein, linking volume regulation to Ca2+ signaling in astrocytes. This knowledge provides new insight into the MLC1 protein function and into the mechanisms controlling ion/water exchanges in the brain, which may help identify possible molecular targets for the treatment of MLC and other pathological conditions caused by astrocyte swelling and brain edema.

Mouse Model of Fragile X Syndrome Analyzed by Quantitative Proteomics: A Comparison of Methods

“…In fact, calcium-binding was another gene ontology term identified by both methods and many of the target proteins have a direct role in calcium homeostasis in the brain. For example, calcium levels are buffered by calbindin-1 ( calb1 ) and the membrane protein MLC1 ( mlc1 ) . Both necab2 and wolframin ( wfs1) are sensors of calcium and have a role in the synaptic release. , Anxa1 is a calcium reservoir and CaMKK2, a calcium/calmodulin-dependent protein kinase, has a possible role in neurite growth .…”

“…For example, calcium levels are buffered by calbindin-1 (calb1) 55 and the membrane protein MLC1 (mlc1). 56 Both necab2 and wolframin (wfs1) are sensors of calcium and have a role in the synaptic release. 57,58 Anxa1 is a calcium reservoir 59 and CaMKK2, a calcium/ calmodulin-dependent protein kinase, has a possible role in neurite growth.…”

Megalencephalic leukoencephalopathy with subcortical cysts: a variant update and review of the literature

“…Many signaling cascades and cellular properties are known to modulate VRAC functioning in different cell types (for review see (Jentsch, 2016)). A recent study has suggested that intracellular calcium might be an intermediate (Brignone et al, 2022), through CaMKII-mediated phosphorylation of MLC1. AQP4 forms a complex with TRPV4 and together these proteins affect RVD through a Frontiers in Genetics frontiersin.org functional interaction with VRAC (Benfenati et al, 2011).…”

Piezo1, the new actor in cell volume regulation

“…In several cell systems, exposure to hypotonic stimuli triggers a rise in intracellular Ca 2+ concentration, as the result of both Ca 2+ influx from extracellular space through MSCs [ 8 , 15 , 19 , 83 , 86 , 96 , 124 , 161 ] and its release from internal stores [ 99 , 127 , 149 ]. Despite the ubiquity of cytosolic Ca 2+ transients evoked by hypotonic cell swelling, intracellular Ca 2+ signals are not always needed for the RVD response.…”