Defining the Role of Erlin2, an ER Membrane Protein of the Erlin1/2 Complex That Mediates Ubiquitination of the Inositol Trisphosphate Receptor (IP3R1)

Natalie Rebeyev, Syracuse University


Ataxias and neurodegenerative diseases affect neurons (nerve cells) in the brain and spinal cord. Those who suffer from spinocerebellar ataxias (SCA) experience degeneration of the spinal cord in the dorsal columns and dorsal column nuclei, and of the cerebellum (located at the base of the brain), which controls coordination and voluntary movement. Abnormal function of cells is followed by their demise. As these processes proceed, patients suffer from an array of symptoms, which include problems with coordination, vision, gait and memory loss. As more neurons die, symptoms worsen, and sadly, this results in an individual’s inability to function in the world.

Autosomal dominant sensory ataxia (ADSA), also known as neuroaxonal dystrophy, a spinocerebellar ataxia with other neurological signs, is caused by degeneration of the posterior columns of the spinal cord. This disease is genetic in origin and like many others, is progressive. Furthermore, it is characterized by cerebellar atrophy linked to complications involving balance, posture, and voluntary muscle movements. ADSA is caused by a mutation in the E3 ubiquitin ligase, RNF170, an enzyme that facilitates recognition and degradation of the inositol 1,4,5-trisphosphate (IP3) receptor through the ubiquitin proteasome pathway. Studying RNF170, and other proteins that associate with the receptor to carry out endoplasmic reticulum associated degradation is important because it can revolutionize advances in trying to treat ADSA.

Other prominent proteins that play a role in receptor degradation are found in the erlin1/2 complex, which serves as a recognition factor, to which RNF70 is constitutively bound. We looked at the basic growth patterns of αT3-1 mouse gondatrope cells in wild type, cells replete with erlin2 and lacking erlin2, and cells replete with RNF170 and lacking RNF170. We investigated the interaction between erlin2 and RNF170 by trying to determine if ubc13 is the E2 that attaches to the receptor, because it is the only known E2 to make lysine-63 chains, by which the receptor is ubiquitinated. This was done through co-immunoprecipitation and through the use of cross-linkers. Furthermore, we looked at using the CRISPR/Cas9 system to knock out ubc13 in the future.

There was no difference in growth between the cell types, even when stimulated by GnRH. There was no clear interaction between ubc13 (E2) and RNF170 (E3) in all of the experiments. We hope to use CRISPR to show that lysine-63 chains are not made when ubc13 is not present, in order to potentially validate that ubc13 is the E2 that binds to RNF170, thereby allowing for degradation of the IP3 receptor.