Integrin-Binding Proteins Kindlins Regulate Kidney Branching Morphogenic in Both Integrin β1-Dependent and -Independent Mechanisms

Background: The kidney collecting system derives from the branching of the ureteric bud (UB). This process is dependent on growth factor (GF) signaling and the interactions between integrins and the extracellular matrix. Integrin function requires binding of structural intracelluar proteins such as kindlins to highly conserved NxxY motifs to the β cytoplasmic tails. The kidney contains two Kindlins, namely Kindlin 1 and 2, yet their role in govering β1-mediated UB branching is currently unknown. Methods: Two mutations in the β1 cytoplasmic tail can abrogate kindlin-β1 binding (TT/AA (amino acids 800-801) and Y795A). We expressed these mutations in the developing mouse UB using the Hoxb7 promoter (Hoxb7:TTAA, and Hoxb7:Y795A) and assessed 3D branching. We compared this phenotype to mice lacking both Kindlin 1 and 2 in the UB (Hoxb7:K1/K2). Furthermore, we generated collecting duct (CD) cells in vitro expressing these mutations or lacking kindlins and assessed tubulogenesis, polarity, adhesion, migration, proliferation, and GF-dependent signaling. Results: Hoxb7:TTAA and Hoxb7:Y795A mice displayed mildly dysplastic collecting systems with a mild branching morphogenesis defect. CD cells expressing TTAA or Y795A mutations had moderate abnormalities in tubulogenesis, cell adhesion, migration, proliferation, and polarity, while matrix-induced and GF-dependent signaling was largely intact. In contrast, mice lacking both kindlins in the UB failed to form kidneys due to a lack of UB budding. In addition, the K1/K2 KO CD cells had severe spreading, and polarity defects, and were unable to adhere or transduce GF signaling. Conclusion: This study shows that kindlins are essential for UB formation and that they control UB branching through mechanisms that are largely independent of β1 integrin binding. Funding: NIDDK Support