Striated muscles contain a tightly ordered cytoplasm in which the shape and size of the nuclei are comparable and nuclear distribution is uniform. These features were recently shown to be essential for muscle function. In an attempt to elucidate mechanisms regulating the position and shape of myonuclei, we analyzed the function of the two KASH proteins that are uniquely present in the Drosophila genome, MSP-300 and Klarsicht, both expressed in striated muscles. We demonstrated that both KASH proteins cooperate to construct a unique ring composed of MSP-300 protein that surrounds and attached to the nuclear envelope. The MSP-300 nuclear ring structure recruits and associates with a network of polarized astral microtubules that enables the dynamic movement and uniform spacing between the nuclei in each muscle fiber.