Gain-of-Function Mutations in KCNN3 Encoding the Small-Conductance Ca2+-Activated K+ Channel SK3 Cause Zimmermann-Laband Syndrome

Abstract

Zimmermann-Laband syndrome (ZLS) is characterized by coarse facial features with gingival enlargement, intellectual disability (ID), hypertrichosis, and hypoplasia or aplasia of nails and terminal phalanges. De novo missense mutations in KCNH1 and KCNK4, encoding K þ channels, have been identified in subjects with ZLS and ZLS-like phenotype, respectively. We report de novo missense variants in KCNN3 in three individuals with typical clinical features of ZLS. KCNN3 (SK3/KCa2.3) constitutes one of three members of the small-conductance Ca 2þ -activated K þ (SK) channels that are part of a multiprotein complex consisting of the pore-forming channel subunits, the constitutively bound Ca 2þ sensor calmodulin, protein kinase CK2, and protein phosphatase 2A. CK2 modulates Ca 2þ sensitivity of the channels by phosphorylating SK-bound calmodulin. Patch-clamp whole-cell recordings of KCNN3 channel-expressing CHO cells demonstrated that disease-associated mutations result in gain of function of the mutant channels, characterized by increased Ca 2þ sensitivity leading to faster and more complete activation of KCNN3 mutant channels. Pretreatment of cells with the CK2 inhibitor 4,5,6,7-tetrabromobenzotriazole revealed basal inhibition of wild-type and mutant KCNN3 channels by CK2. Analogous experiments with the KCNN3 p.Val450Leu mutant previously identified in a family with portal hypertension indicated basal constitutive channel activity and thus a different gain-of-function mechanism compared to the ZLS-associated mutant channels. With the report on de novo KCNK4 mutations in subjects with facial dysmorphism, hypertrichosis, epilepsy, ID, and gingival overgrowth, we propose to combine the phenotypes caused by mutations in KCNH1, KCNK4, and KCNN3 in a group of neurological potassium channelopathies caused by an increase in K þ conductance.

Deciphering the molecular mechanism of SK2 channel activation by intracellular calcium to develop new therapeutic agents

“…1 Regarding the consequences of in vivo mutations, a rare S436C mutation in SK3 channels leads to a disease called the Zimmermann-Laband syndrome affecting the development of many types of tissue such as gums, nails and facial features. 19 This mutation located at the interface between calmodulin and one of the channel's subunits induces a faster and more complete activation of the corresponding channel similarly to what is observed in the Nam's paper. Indeed, the replacement of the serine 436 by a cysteine 19 induces an increase of the local hydrophobicity and reduced propensity to create hydrogen bond leading to structural changes also affecting the gating.…”

Dyshomeostatic modulation of Ca2+-activated K+ channels in a human neuronal model of KCNQ2 encephalopathy

“…Although, increased K + conductance may not be traditionally associated with hyperexcitability and epilepsy, several studies have shown that inhibition of K + conductances improves learning and memory in wild-type mice, or seizure susceptibility in a model of Angelman syndrome ( Stackman et al, 2002 ; Fontán-Lozano et al, 2011 ; Sun et al, 2019 ). Furthermore, gain-of-function mutations in several K + channels including KCNQ2, KCNQ3, KCNQ5, BK, SK3, KCNT1, and K V 4.2 have been reported in genetic epilepsy and developmental disorders ( Niday and Tzingounis, 2018 ; Bauer et al, 2019 ). Because KCNQ2-DEE patients present with seizures in the first days of life, targeting the underlying cause might not be effective if started late in the disease course.…”

Hydrophobic interactions between the HA helix and S4‐S5 linker modulate apparent Ca2+ sensitivity of SK2 channels

“…46 Mutation of the equivalent serine in SK3 channels to cysteine (S436C) also increases apparent Ca 2+ sensitivity. 47 This CaM-S 45 A interaction interface was then explored as potential binding pocket of small molecule modulators of the channels. 46 Now that we know that CaM interactions with both the CaMBD (HA and HB helices) and the S4-S5 linker (S 45 A and S 45 B helices) are crucial for the apparent Ca 2+ sensitivity of the channels, then what about the interactions between the CaMBD and the S4-S5 linker?…”