Studying the biochemical basis for the action of ABERRANT LATERAL ROOT FORMATION4, a putative ubiquitylation tuner

• Applicant: Professor Dr. Steffen Abel, since 7/2022
• Subject Area: Plant Biochemistry and Biophysics
• Term: from 2019 to 2023
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 421703589

Ubiquitin-mediated proteolysis is the ultimate universal cellular switch controlling almost every aspect of growth and development. A hierarchical ATP-dependent E1-E2-E3 enzymatic cascade facilitates specific protein ubiquitylation. Modular Cullin 1–RING E3 ubiquitin ligases (a.k.a SCFs) control a plethora of biological processes, and are subjected to precise tuning at the level of complex assembly, target recruitment, and Ub transfer to the targets. Despite the profound impact of ubiquitylation on proteostasis, we know little about how the catalysis and regulation of SCFs and the initial Ub transfer event on protein targets takes place. Glomulin (GLMN)/ABERRANT LATERAL ROOT FORMATION4 (ALF4) is a single copy gene across kingdoms, and the GLMN/ALF4 protein inhibits SCF activity, as it binds and competes for the E2 docking site. Knockout glmn/alf4 mutants are lethal in both mammals and plants, emphasizing GLMN/ALF4’s essential function. Mutants expressing weak alf4 alleles in Arabidopsis, although viable, show growth phenotypes that suggest shortcomings in plant hormone responses. We previously showed that defects in ALF4 destabilize components of SCFs, thereby causing stabilization of protein targets that undergo ubiquitylation in response to phytohormones. Currently, we lack information about why, how, and when ALF4 exerts its inhibitory effect on SCF-E3 ligases and target ubiquitylation. In this proposal, we plan to implement biochemical and biophysical approaches to mechanistically understand ALF4 action. We seek to dissect the effect of ALF4 during ubiquitylation of SCF targets in vivo and in vitro and assess the ALF4-targeted proteome. Also, by building up an ALF4 interaction network we strive to pinpoint how ALF4 activity is regulated. With our research, we certainly will get a better understanding of E3s assembly, and ubiquitin-mediated turnover of proteins.

• DFG Programme: Research Grants
• Ehemalige Antragstellerin: Dr. Luz Irina Calderón Villalobos, until 7/2022