Titin, the largest protein known to date, has been linked to sarcomere assembly 
and function through its elastic adaptor and signaling domains. Titin's M-line 
region contains a unique kinase domain that has been proposed to regulate 
sarcomere assembly via its substrate titin cap (T-cap). In this study, we use a 
titin M line-deficient mouse to show that the initial assembly of the sarcomere 
does not depend on titin's M-line region or the phosphorylation of T-cap by the 
titin kinase. Rather, titin's M-line region is required to form a continuous 
titin filament and to provide mechanical stability of the embryonic sarcomere. 
Even without titin integrating into the M band, sarcomeres show proper spacing 
and alignment of Z discs and M bands but fail to grow laterally and ultimately 
disassemble. The comparison of disassembly in the developing and mature knockout 
sarcomere suggests diverse functions for titin's M line in embryonic development 
and the adult heart that not only involve the differential expression of titin 
isoforms but also of titin-binding proteins.