TY - JOUR
T1 - Membrane distribution of the glycine receptor alpha 3 studied by optical super-resolution microscopy
AU - Notelaers, Kristof
AU - Rocha, Susana
AU - Paesen, Rik
AU - Swinnen, Nina
AU - Vangindertael, Jeroen
AU - Meier, Jochen C.
AU - Rigo, Jean-Michel
AU - Ameloot, Marcel
AU - Hofkens, Johan
PY - 2014/7
Y1 - 2014/7
N2 - In this study, the effect of glycine receptor (GlyR) ?3 alternative RNA splicing on the distribution of receptors in the membrane of human embryonic kidney 293 cells is investigated using optical super-resolution microscopy. Direct stochastic optical reconstruction microscopy is used to image both ?3K and ?3L splice variants individually and together using single- and dual-color imaging. Pair correlation analysis is used to extract quantitative measures from the resulting images. Autocorrelation analysis of the individually expressed variants reveals clustering of both variants, yet with differing properties. The cluster size is increased for ?3L compared to ?3K (mean radius 92 ? 4 and 56 ? 3 nm, respectively), yet an even bigger difference is found in the cluster density (9,870 ? 1,433 and 1,747 ? 200 ?m(-2), respectively). Furthermore, cross-correlation analysis revealed that upon co-expression, clusters colocalize on the same spatial scales as for individually expressed receptors (mean co-cluster radius 94 ? 6 nm). These results demonstrate that RNA splicing determines GlyR ?3 membrane distribution, which has consequences for neuronal GlyR physiology and function.
AB - In this study, the effect of glycine receptor (GlyR) ?3 alternative RNA splicing on the distribution of receptors in the membrane of human embryonic kidney 293 cells is investigated using optical super-resolution microscopy. Direct stochastic optical reconstruction microscopy is used to image both ?3K and ?3L splice variants individually and together using single- and dual-color imaging. Pair correlation analysis is used to extract quantitative measures from the resulting images. Autocorrelation analysis of the individually expressed variants reveals clustering of both variants, yet with differing properties. The cluster size is increased for ?3L compared to ?3K (mean radius 92 ? 4 and 56 ? 3 nm, respectively), yet an even bigger difference is found in the cluster density (9,870 ? 1,433 and 1,747 ? 200 ?m(-2), respectively). Furthermore, cross-correlation analysis revealed that upon co-expression, clusters colocalize on the same spatial scales as for individually expressed receptors (mean co-cluster radius 94 ? 6 nm). These results demonstrate that RNA splicing determines GlyR ?3 membrane distribution, which has consequences for neuronal GlyR physiology and function.
KW - Super-resolution microscopy
KW - Direct stochastic optical reconstruction microscopy
KW - Pair correlation analysis
KW - Glycine receptor
KW - alpha 3 Subunit
KW - RNA splicing
U2 - 10.1007/s00418-014-1197-y
DO - 10.1007/s00418-014-1197-y
M3 - Article
C2 - 24553792
SN - 0948-6143
VL - 142
SP - 79
EP - 90
JO - Histochemistry and Cell Biology
JF - Histochemistry and Cell Biology
IS - 1
ER -