The southern terminus of the highway is at an intersection with U.S. Route 72 (US 72) at Paint Rock.

The northern terminus of the highway is at the Tennessee state line, north of Francisco.

SR 65 and Tennessee State Route 97 serve as a connecting route between US 72 in northern Alabama and US 64 in Franklin County, Tennessee.

From SR 65's southern terminus, the highway is aligned along a two-lane road with numerous curves as it travels through the mountainous regions of northeastern Alabama and southern Tennessee.

North of Paint Rock, SR 65 travels through the unincorporated communities of Garth, Trenton, Hollytree, Princeton, Swaim, and Francisco, before it crosses into Tennessee.