Dataset Information

Allostery between two binding sites in the ion channel subunit TRIP8b confers binding specificity to HCN channels.

ABSTRACT: Tetratricopeptide repeat (TPR) domains are ubiquitous structural motifs that mediate protein-protein interactions. For example, the TPR domains in the peroxisomal import receptor PEX5 enable binding to a range of type 1 peroxisomal targeting signal motifs. A homolog of PEX5, tetratricopeptide repeat-containing Rab8b-interacting protein (TRIP8b), binds to and functions as an auxiliary subunit of hyperpolarization-activated cyclic nucleotide (HCN)-gated channels. Given the similarity between TRIP8b and PEX5, this difference in function raises the question of what mechanism accounts for their binding specificity. In this report, we found that the cyclic nucleotide-binding domain and the C terminus of the HCN channel are critical for conferring specificity to TRIP8b binding. We show that TRIP8b binds the HCN cyclic nucleotide-binding domain through a 37-residue domain and the HCN C terminus through the TPR domains. Using a combination of fluorescence polarization- and co-immunoprecipitation-based assays, we establish that binding at either site increases affinity at the other. Thus, allosteric coupling of the TRIP8b TPR domains both promotes binding to HCN channels and limits binding to type 1 peroxisomal targeting signal substrates. These results raise the possibility that other TPR domains may be similarly influenced by allosteric mechanisms as a general feature of protein-protein interactions.

SUBMITTER: Lyman KA

PROVIDER: S-EPMC5663874 | biostudies-other | 2017 Oct

REPOSITORIES: biostudies-other

ACCESS DATA

Publications

Allostery between two binding sites in the ion channel subunit TRIP8b confers binding specificity to HCN channels.

Lyman Kyle A KA Han Ye Y Heuermann Robert J RJ Cheng Xiangying X Kurz Jonathan E JE Lyman Reagan E RE Van Veldhoven Paul P PP Chetkovich Dane M DM

The Journal of biological chemistry 20170908 43

Tetratricopeptide repeat (TPR) domains are ubiquitous structural motifs that mediate protein-protein interactions. For example, the TPR domains in the peroxisomal import receptor PEX5 enable binding to a range of type 1 peroxisomal targeting signal motifs. A homolog of PEX5, tetratricopeptide repeat-containing Rab8b-interacting protein (TRIP8b), binds to and functions as an auxiliary subunit of hyperpolarization-activated cyclic nucleotide (HCN)-gated channels. Given the similarity between TRIP8 ...[more]

PMID: 28887304

Similar Datasets

Project description:Although hyperpolarization-activated cation (HCN) ion channels are well established to underlie cardiac pacemaker activity, their role in smooth muscle organs remains controversial. HCN-expressing cells are localized to renal pelvic smooth muscle (RPSM) pacemaker tissues of the murine upper urinary tract and HCN channel conductance is required for peristalsis. To date, however, the Ih pacemaker current conducted by HCN channels has never been detected in these cells, raising questions on the identity of RPSM pacemakers. Indeed, the RPSM pacemaker mechanisms of the unique multicalyceal upper urinary tract exhibited by humans remains unknown. Here, we developed immunopanning purification protocols and demonstrate that 96% of isolated HCN+ cells exhibit Ih . Single-molecule STORM to whole-tissue imaging showed HCN+ cells express single HCN channels on their plasma membrane and integrate into the muscular syncytium. By contrast, PDGFR-α+ cells exhibiting the morphology of ICC gut pacemakers were shown to be vascular mural cells. Translational studies in the homologous human and porcine multicalyceal upper urinary tracts showed that contractions and pacemaker depolarizations originate in proximal calyceal RPSM. Critically, HCN+ cells were shown to integrate into calyceal RPSM pacemaker tissues, and HCN channel block abolished electrical pacemaker activity and peristalsis of the multicalyceal upper urinary tract. Cumulatively, these studies demonstrate that HCN ion channels play a broad, evolutionarily conserved pacemaker role in both cardiac and smooth muscle organs and have implications for channelopathies as putative aetiologies of smooth muscle disorders. KEY POINTS: Pacemakers trigger contractions of involuntary muscles. Hyperpolarization-activated cation (HCN) ion channels underpin cardiac pacemaker activity, but their role in smooth muscle organs remains controversial. Renal pelvic smooth muscle (RPSM) pacemakers trigger contractions that propel waste away from the kidney. HCN+ cells localize to murine RPSM pacemaker tissue and HCN channel conductance is required for peristalsis. The HCN (Ih ) current has never been detected in RPSM cells, raising doubt whether HCN+ cells are bona fide pacemakers. Moreover, the pacemaker mechanisms of the unique multicalyceal RPSM of higher order mammals remains unknown. In total, 97% of purified HCN+ RPSM cells exhibit Ih . HCN+ cells integrate into the RPSM musculature, and pacemaker tissue peristalsis is dependent on HCN channels. Translational studies in human and swine demonstrate HCN channels are conserved in the multicalyceal RPSM and that HCN channels underlie pacemaker activity that drives peristalsis. These studies provide insight into putative channelopathies that can underlie smooth muscle dysfunction.

Dataset Information

Allostery between two binding sites in the ion channel subunit TRIP8b confers binding specificity to HCN channels.

Publications

Allostery between two binding sites in the ion channel subunit TRIP8b confers binding specificity to HCN channels.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets