To determine whether PDGFR-α/β is regulated by miR-34a through direct binding to the 3′ UTR of PDGFR-α/β, we constructed a series of 3′ UTR fragments, including the full-length wild-type PDGFR-α 3′ UTR, a binding site 1 mutant, a binding site 2 mutant, the full-length wild-type PDGFR-β3′ UTR, and a PDGFR-β binding site mutant.
But the finding that physiological changes in blood pressure show the same direction in KO and G2019S mutants could suggest that the net output of LRRK2 signaling by the G2019S mutant and the full KO might be similar.
Upon physical removal of the spore coat, the mutant lacking the full gerP operon no longer exhibited a germination defect, suggesting that the GerP proteins play a role in spore coat permeability.
A similar strategy was used to create the lsk1-K306R GFP tagged mutant, only the full length product was cloned into the EcoRI and SmaI sites of the pJK210-GFP vector.
The G576E mutation would exert both charge and space hindrances on the side chain of L574 and thus disrupt the dimerization (Figure 7), consistent with the previous observation that the G576E mutant of the full length GNE remains as a trimer [11].
Although R97 was involved in over 80% of the induced NF-κB, the R97Q mutant generated the full A20 transcript potential.
We then concentrated on PKCζ, first generating a mutant of the full length construct which mimics the PKCι situation (aPKCζ G141R).
To construct the eIF4E mutants, the full-length eIF4E coding sequence was cloned into entry vector pDONR207 by recombination reactions using BP clonase II (Invitrogen).
First, the expression levels of all TRAF2 mutants, including the full length proteins were analyzed, and then the cell lysates with equal expression of TRAF2 mutant proteins were immunoprecipitated using anti-FLAG antibody.
Results showed specific Isl1-Cre expression in mutants displaying the full spectrum of abnormalities (supplementary material Fig. S2C–E').
