Figure EV4.

WNK4 and SPAK puncta do not colocalise with LAMP1 or form ubiquitin-containing aggregates

Representative pseudocoloured confocal single focal plane images of immunofluorescently stained kidney sections (n = 4 per genotype). WNK4 and SPAK form discrete puncta in the distal convoluted tubule of CUL3^{WT/Δ403–459} mice. These puncta do not colocalise with lysosomes (LAMP1) or form ubiquitylated aggregates. The puncta have a predominantly basolateral preference with several large juxta-nuclear puncta per cell.

1. Immunolocalisation of total SPAK protein (tSPAK) puncta and LAMP1 in the distal convoluted tubule of CUL3^{WT/Δ403–459} versus CUL3^WT mice at a 4-h fasting baseline. Scale bar, 20 μm.
2. Immunolocalisation of total WNK4 puncta, ubiquitin and LAMP1 in the distal convoluted tubule of CUL3^{WT/Δ403–459} versus CUL3^WT mice at a 4-h fasting baseline. Scale bar, 10 μm.