ATXN1 interacts with FOX-2 splice variants.

<p>(<b>A</b>) Schematic view of ATXN1 and the regions used in the Y2H analyses (left) and FOX-2 variants (right). Prey plasmids pACT-ATXN1-NTQ30 or pACT-ATXN1-NTQ82 cover amino acids 1–576, pACT-ATXN1-AXH amino acids 559–701, and pACT-ATXN1-CT amino acids 530–816. Bait plasmid pBTM-FOX-2_V1 covers amino acids 1–380 of variant 1, including the putative NLS in the C-terminal region, and pBTM-FOX-2_cyt covers amino acids 1–391 of the cytoplasmic FOX-2 variant. Different C-terminal regions of both FOX-2 variants are highlighted in blue and green. (<b>B; C</b>) For directed Y2H analyses yeast strain L40ccua was co-transformed with the relevant bait and prey plasmids, and transformants were selected and spotted onto selective media or membrane to analyze activity of the reporter genes. (<b>D</b>) For Co-IP experiments, HEK293T cell lysates derived from cells overexpressing FLAG-ATXN1-Q30 (left panel) or HEK293T cell lysates (right panel) were incubated with an antibody directed against FOX-2 (Bethyl or Abnova, respectively). Cell lysates incubated without primary antibody served as controls. Then, membranes were treated with an antibody directed against the FLAG tag or ATXN1 to detect precipitated protein.