ed, when rat kidney sections examined body with an anti-PP2A A subunit-antibody. Immunpr Zipitation of kidney tissue Immunpr Zipitation was conducted from kidney tissue, whether the Na, K-ATPase associated with PP2A to determine in situ. Immunpr Zipitationen were addressed GDC-0941 957054-30-7 with an antique Rpern aortic PP2A C-subunit and an antique Body against the HA epitope, as controlled by out Negative. Na, K-ATPase subunit, the employees and found working with Lltem PP2A by Western blotting with biotinylated anti-Na, K-ATPase subunit Antique Body was. The biotinylated Na, K-ATPase subunit antibody Body was used to the need for a secondary Ren Antique Body, which also recognize k Can Antik Body, which for the Immunpr were Zipitation used to avoid.
If an antique Body for the team of professionals Immunpr Zipitation was used, Belinostat PX105684 a band migrating with the extremely low CO-Na, K-ATPase subunit was detected. However, both the A and anti-PP2A subunit are antique C body clearly executed Co filled easily detectable amounts of Na, K-ATPase subunit. The amount of the subunit was below was gr It in the PP2A A subunit-antibody Body against the Antique Body when subunit C was used drawn. This difference k Nnte to the different accessibility of the antigenic site responsible for the PP2A A or Csubunit Antique Body complex in the Na, K-situ ATPase/PP2A. In Similar way can PP2A A and C subunits of antique Rpern against different affinity Th for their respective antigens. In both cases Fill this result best CONFIRMS the conclusion that the Na, K-ATPase in situ associated with PP2A.
Characterization of the interaction of Na, K-ATPase subunit and PP2A C and A subunits We investigated the dependence Dependence of the interaction between the ATPase Na, K and PP2A on the expression of PP2A A or C units for these experiments were COS cells with cDNA HA Co or flag labeled subunits of PP2A and a cDNA encoding Chim re H85N-subunit construct transfected. H85N is a Chim Re, in which the first 85 residues of the Na, K-ATPase subunit of the enzyme which gastric H, K-ATPase replaced. This Chim Re obvious functional properties that are recognized identical to those of ATPase Na, K, and is carried from the Antique Body against HK9 the N-terminus of the H, K-ATPase asubunit directed. Fig. 3A, the development of Western blot of transfected COS cell lysates shows subjected Immunpr Zipitation with antique Body HK9 then rpern HA antibody That Recogn t detects the C subunit exogenous PP2A.
As expected, when the cells were transfected with HA-subunit C, was very few Csubunit PP2A in Immunpr Zipitat HK9 observed. However, we found that PP2A subunit C immunpr Was zipitiert when H85N was with PP2A C subunit co-expressed PP2A C-subunit was in Immunpr Zipitaten HK9 when the cells were detected in 1. Immunolocalization of Na, K-ATPase and PP2A in situ. Mouse kidney sections were stained with Na, K-ATPase Antique Body and Antique Body-PP2A subunit C Rbt Merged images are shown in C Na, K-ATPase and PP2A partially come together in the folds of the basolateral proximal Tubul Ren epithelial cells. Typical results showed one of three experiments. doi: 10.1371/journal.pone.0029269.
g001 interaction between PP2A and the Na, K-ATPase PLoS ONE | Published in PloSOne second December 2011 | Volume 6 | Issue 12 | e29269 H85N and transfected with both subunits A and C PP2A. PP2A A subunit has no apparent effect on activation or inhibition of the interaction between the C-subunit and PP2A Na, K-ATPase subunit. Figure 3b shows Immunpr Zipitation cooperation H85N and the flag of the subunit. Again, very few Asubunit PP2A in HK9 Immunpr was Zipitation detected when cells were transfected with PP2A A subunit alone. PP2A A subunit was H85N with time in the absence and presence of exogenous C-subunit of PP2A immunpr Zipitiert. The interaction between PP2A and Asubunit H85N was reduced approximately in the presence of an excess of PP2A subunit C These results show that the Na, K-ATPase subunit forms a complex with both of the exogenous