Functional Characterization of Two E3 Ubiquitin Ligases (PUB2 and PUB4) and the EXTRA LARGE G PROTEINS in Arabidopsis

As sessile organisms, plants need to adjust their growth and development according to ever-changing environments. Environmental cues and endogenous signals are perceived by receptor or sensor proteins and transduced through sophisticated signaling pathways into appropriate cellular responses. Various G proteins and protein kinases are among the most common signal transduction components, and protein ubiquitination is also a prominent mechanism that regulates protein function and cell signaling. The signaling pathways often lead to transcriptional reprogramming that facilitates physiological and developmental response.

We initially identified Arabidopsis EXTRA LARGE G PROTEIN 2 (XLG2) and XLG3 as positive regulators of immunity, which are now known to form heterotrimeric G protein complexes. Later, we identified two E3 ubiquitin ligases, PUB2 (PLANT U-BOX PROTEIN 2) and PUB4, as XLG-interacting proteins. The xlg and pub mutants, including their double and triple mutants, share similar pleiotropic phenotypes, such as defects in cell expansion, proliferation, and programmed cell death as well as in hormone response. The findings from our study and by other groups have indicated that these PUB and XLG proteins are newly discovered players in multiple important developmental and physiological pathways.

The main goal of this proposal is to define the functions of the PUB and XLG proteins and their modes of action in mediating those cellular processes. Through genetic and molecular approaches, we have identified additional genes/proteins that physically and/or functionally interact with PUB4 and their roles in the PUB/XLG-mediated processes will be affirmed and further defined. We will determine whether PUB2/PUB4 regulate functions of their candidate substrates (including XLGs) by mono- ubiquitination. The molecular function of PUB4 in NITRILASE1-catalyzed auxin biosynthesis will be elucidated. The effects of the pub and xlg mutations on the cytokinin response will be investigated. By understanding the mechanisms by which these PUB and XLG proteins function, the proposed study will offer novel insights into the roles of protein ubiquitination and G proteins in regulating important cellular processes.