Structural basis for the dominant or recessive character of GLIALCAM mutations found in leukodystrophies

Abstract

Megalencephalic leukoencephalopathy with subcortical cysts (MLC) is a type of leukodystrophy characterized by white matter edema, and it is caused mainly by recessive mutations in MLC1 and GLIALCAM genes. These variants are called MLC1 and MLC2A with both types of patients sharing the same clinical phenotype. In addition, dominant mutations in GLIALCAM have also been identified in a subtype of MLC patients with a remitting phenotype. This variant has been named MLC2B. GLIALCAM encodes for an adhesion protein containing two immunoglobulin (Ig) domains and it is needed for MLC1 targeting to astrocyte–astrocyte junctions. Most mutations identified in GLIALCAM abolish GlialCAM targeting to junctions. However, it is unclear why some mutations behave as recessive or dominant. Here, we used a combination of biochemistry methods with a new developed anti-GlialCAM nanobody, double-mutants and cysteine cross-links experiments, together with computer docking, to create a structural model of GlialCAM homo-interactions. Using this model, we suggest that dominant mutations affect different GlialCAM–GlialCAM interacting surfaces in the first Ig domain, which can occur between GlialCAM molecules present in the same cell (cis) or present in neighbouring cells (trans). Our results provide a framework that can be used to understand the molecular basis of pathogenesis of all identified GLIALCAM mutations.

Control of membrane protein homeostasis by a chaperone-like glial cell adhesion molecule at multiple subcellular locations

“…The data that GlialCAM disease R92W mutation in the N-terminal IgV adhesion domain has reduced PM stability agree with a requirement of extracellular adhesion of GlialCAM to position MLC1 at the PM 27 , 55 . GlialCAM partitions the PM signalling cluster and the misaligned transmembrane phase separation seems to contribute to interfered interactions of signalling complexes in GlialCAM expressing cells both in trans and cis 55 . Notably, the neuroplastin CAMs have a similar anchoring role for the GluR1, GABA A , and monocarboxylate transporters 56 , 57 .…”

Megalencephalic Leukoencephalopathy with Subcortical Cysts Disease-Linked MLC1 Protein Favors Gap-Junction Intercellular Communication by Regulating Connexin 43 Trafficking in Astrocytes

“…In human and mouse brains, the MLC1/GlialCAM protein complex is highly expressed at perivascular astrocyte end-feet contacting the BBB and is particularly enriched at astrocyte–astrocyte junctions, where it establishes interactions with ZO-1 and Cx43 [ 24 , 40 ]. The diffuse brain edema characterizing the magnetic resonance imaging (MRI) of MLC patients and astrocyte vacuolation/swelling observed by histological analysis of brain tissue (see Reference [ 46 ] and the reference therein) along with alterations of chloride, potassium, and calcium fluxes reported in MLC disease models [ 27 , 48 , 49 , 50 ] suggest that intercellular communication and glial syncytium properties might be affected directly or indirectly by MLC1 mutations. A further support to this hypothesis is provided by the finding that in astrocytoma cells GlialCAM pathological mutations disrupt GlialCAM/Cx43 interactions, decreasing Cx43 stability at the plasma membrane [ 40 ].…”

“…The development of possible therapeutic interventions is hampered by the lack of knowledge about MLC molecular pathogenesis and the proper function of MLC1, the protein, found mutated in 80% of patients. The discovery of a second protein linked to MLC, the MLC1 molecular interactor GlialCAM [21], allowed identifying two disease variants, MLC2A and B, caused by recessive and dominant mutations in GlialCAM that show some differences in the disease progression [19,46,47]. With the aim of shedding light on MLC molecular pathogenesis, we here investigated Cx43 properties in U251 astrocytoma cell lines overexpressing MLC1 WT or carrying pathological mutations.…”

Megalencephalic leukoencephalopathy with subcortical cysts 1 (MLC1) promotes glioblastoma cell invasion in the brain microenvironment

“…In conjunction with Mlc1, other interacting proteins such as GlialCAM are likely involved in promoting GSC polarity and invasion. In astrocytes, post-translational trafficking of Mlc1 from the endoplasmic reticulum to the cell membrane is dependent upon interactions with GlialCAM [ 47 ]. Mlc1/GlialCAM interact with Aqp4 and Trpv4, both components of the dystrophin-glycoprotein complex in astrocytes which also includes the intracellular adapter proteins α-dystrobrevin and dystrophin, α/β-dystroglycan transmembrane proteins, and matrix proteins such as agrin, laminin, and integrin adhesion.…”