Tropomodulin, a tropomyosin-dependent actin filament capping protein, consists of two structurally and functionally different domains. Tropomodulin lacking its C-terminal domain can cap actin filaments with an affinity close to the full-length protein in vitro. To investigate the functional properties of the C-terminal domain in live cells, truncated GFP-tagged tropomodulin was expressed in rat cardiac myocytes. Three fragments were analyzed: Tmod1(1-159) that lacks the entire C-terminal domain, Tmod1(1-320) and Tmod1(1-349). GFP-Tmod1(1-159) did not assemble well at the pointed ends of the filaments (∼80% of the cells demonstrated a diffuse distribution, while ∼20% showed faint, inconsistent assembly). Together, the in vitro and live cell studies indicate that the C-terminal domain is not required for capping actin filaments but is important for specifically targeting tropomodulin to the pointed ends of the actin filaments in sarcomeres. In the cells where GFP-Tmod1(1-320) was expressed most (∼70%) of the assembly was faint and inconsistent. Tmod1(1-320) lacks the 39 C-terminal residues that include both the C-terminal helix that is not a part of LRR fold, and the tropomyosin independent actin filament capping site. The precise location of the tropomyosin independent actin filament capping site is not known, although removal of 15 residues from the C-terminus destroys the actin filament capping ability of Tmod1 in the absence of tropomyosin. GFP-Tmod1(1-349) is missing the ten C-terminal residues which discriminates Tmod1 from other tropomodulin isoforms; this fragment consistently assembled at the thin filament pointed ends, comparable to the cells expressing wild type GFP-Tmod1. Based on these data we suggest that both the LRR fold (residues 160-320) and residues 321-349 are important for regulating tropomodulin's pointed end capping activity though the specific role each of these regions play in this phenomenon may be different.