Glutamate receptors at the postsynaptic side translate neurotransmitter release from presynapses into postsynaptic excitation. They play a role in many forms of synaptic plasticity, e.g., homeostatic scaling of the receptor field, activity-dependent synaptic plasticity and the induction of presynaptic homeostatic potentiation (PHP). The latter process has been extensively studied at Drosophila melanogaster neuromuscular junctions (NMJs). The genetic removal of the glutamate receptor subunit IIA (GluRIIA) leads to an induction of PHP at the synapse. So far, mostly imprecise knockouts of the GluRIIA gene have been utilized. Furthermore, mutated and tagged versions of GluRIIA have been examined in the past, but most of these constructs were not expressed under endogenous regulatory control or involved the mentioned imprecise GluRIIA knockouts. We performed CRISPR/Cas9-assisted gene editing at the endogenous locus of GluRIIA. This enabled the investigation of the endogenous expression pattern of GluRIIA using tagged constructs with an EGFP and an ALFA tag for super-resolution immunofluorescence imaging, including structured illumination microscopy (SIM) and direct stochastic optical reconstruction microscopy (dSTORM). All GluRIIA constructs exhibited full functionality and PHP could be induced by philanthotoxin at control levels. By applying hierarchical clustering algorithms to analyze the dSTORM data, we detected postsynaptic receptor cluster areas of ~0.15 µm². Consequently, our constructs are suitable for ultrastructural analyses of GluRIIA.

X-linked recessive (XR) complete MCTS1 deficiency underlies Mendelian susceptibility to mycobacterial disease (MSMD) in patients with bacille Calmette-Guérin (BCG) disease. We investigated the genotypic and phenotypic landscape of four new unrelated families from four distinct countries. Three patients had adverse reactions to the BCG vaccine, whereas another patient was not vaccinated with BCG and had an infection with Mycobacterium abscessus at 16 years of age. Whole-exome sequencing of the probands revealed hemizygosity for rare germline MCTS1 variants. In addition to a previously reported loss-of-expression (LOE) and loss-of-function (LOF) variant, we identified three new MCTS1 variants. The p.L170* and E60Kfs5* variants are LOF, whereas p.W175* is hypomorphic when overexpressed. Thus, we report four new MSMD patients with complete or partial forms of XR MCTS1 deficiency, including three patients with newly discovered genotypes. A diagnosis of partial or complete XR MCTS1 deficiency should be considered in boys and men with MSMD displaying mycobacterial infection.