Endocrine Abstracts

Evidences for a functional link between insulin receptor (IR) and β₂-adrenergic receptor (β₂AR) exist in the mouse heart and in retina and are correlated with the cardiovascular dysfunction under insulin-resistant states¹ and a diabetic retinopathy phenotype². In our recent study we showed that i) IR coexpression reduced β₂AR surface expression and accelerated its internalization, ii) provided evidence for direct interaction and higher-order β₂AR:IR oligomer formation, likely comprising heteromers of homodimers, and iii) identified prospective intracellular interaction domains engaged in heteromerisation³. In this study we examined the involvement of the cytoplasmic part of the IR β chain in heteromerisation and addressed the role of β₂AR:IR heteromer in signalling. Evidence suggesting that the cytoplasmic part of the IR β chain is prerequisite for the interaction with the β₂AR was provided by BRET² saturation and Heteromer Identification Technology (HIT) assays using a IR 1–1271 mutant lacking the IR C-terminal tail region (amino acids from 1272 to 1360). In BRET² saturation assays, IR 1–1271–Rluc8 mutant displayed lower BRET_max value and slightly reduced affinity compared to the WT IR. For the complex consisting of IR 1–1271–RLuc8:β₂AR-GFP² the saturation was not reached; most likely reflecting only random collisions. Furthermore, in the IR 1–1271–Rluc8:β₂AR HIT assay with β-arrestin 2 no substantial agonist-induced increase in BRET² signal was detected that would be indicative of β-arrestin 2 recruitment to the IR 1–1271–Rluc8:β₂AR heteromer, arguing against ability of IR 1–1271 to heteromerise with the β₂AR. Additional in silico mutation analysis of IR predicted substitutions mostly affecting the binding to the β₂AR. β₂AR:IR heteromerization also influenced the pharmacological phenotype of the β₂AR i.e. its ability to recruit β-arrestin 2 and cAMP signalling.

References: 1. Fu Q, Xu B, Liu Y, Parikh D, Li J, et al. Diabetes 2013 63 2676–2689.

2. Jiang Y, Zhang Q, Liu L, Tang J, Kern TS & Steinle JJ. PLoS One 2013 8 (7) e70555.

3. Mandić M, Drinovec L, Glisic S, Veljkovic N, Nøhr J & Vrecl M. PLoS One 2014 9 (11) e112664.

Disclosure: We acknowledge funding from the Basileus program to M Mandic, Slovenian Research Agency program P4-0053 to M Vrecl and the Ministry of Education, Science and Technological Development of the Republic of Serbia (grant number 173001) to S Glisic.