C terminus-mediated control of voltage and cAMP gating of hyperpolarization-activated cyclic nucleotide-gated channels

The hyperpolarization-activated cyclic nucleotide-gated (RON) family of "pa cemaker" channels includes 4 isoforms, the kinetics and cAMP-induced modula tion of which differ quantitatively. Because HCN isoforms are highly homolo gous. in the central region, but diverge more substantially in the N and C termini, we asked whether these latter regions could contribute to the dete rmination of channel properties. To this aim, we analyzed activation/deacti vation. kinetics and the response to cAMP of heterologously expressed isofo rms mHCN1 and rbHCN4 and verified that mHCN1 has much faster kinetics and l ower cAMP sensitivity than rbHCN4. We then constructed rbHCN4 chimeras by r eplacing either the N or the C terminus, or both, with the analogous domain s from mHCN1. We found that: 1) replacement of the N terminus (chimera N1-4 ) did not substantially modify either the kinetics or cAMP dependence of wi ldtype channels; 2) replacement of the C terminus, on the contrary, resulte d in a chimeric channel (4-Cl), the kinetics of which were strongly acceler ated compared with rbHCN4, and that was fully insensitive to cAMP; 3) repla cement of both N and C termini led to the same results as replacement of th e C terminus alone. These results indicate that the C terminus of rbHCN4 co ntributes to the regulation of voltage- and cAMP-dependent channel gating, possibly through interaction with other intracellular regions not belonging to the N terminus.