SUN1 and SUN2 play critical but partially redundant roles in anchoring nuclei in skeletal muscle cells in mice

Abstract

How the nuclei in mammalian skeletal muscle fibers properly position themselves relative to the cell body is an interesting and important cell biology question. In the syncytial skeletal muscle cells, more than 100 nuclei are evenly distributed at the periphery of each cell, with 3-8 nuclei anchored beneath the neuromuscular junction (NMJ). Our previous studies revealed that the KASH domain-containing Syne-1/Nesprin-1 protein plays an essential role in anchoring both synaptic and nonsynaptic myonuclei in mice. SUN domain-containing proteins (SUN proteins) have been shown to interact with KASH domain-containing proteins (KASH proteins) at the nuclear envelope (NE), but their roles in nuclear positioning in mice are unknown. Here we show that the synaptic nuclear anchorage is partially perturbed in Sun1, but not in Sun2, knockout mice. Disruption of 3 or all 4 Sun1/2 wild-type alleles revealed a gene dosage effect on synaptic nuclear anchorage. The organization of nonsynaptic nuclei is disrupted in Sun1/2 doubleknockout (DKO) mice as well. We further show that the localization of Syne-1 to the NE of muscle cells is disrupted in Sun1/2 DKO mice. These results clearly indicate that SUN1 and SUN2 function critically in skeletal muscle cells for Syne-1 localization at the NE, which is essential for proper myonuclear positioning.KASH domain ͉ Nesprin ͉ neuromuscular junction ͉ nuclear envelope protein ͉ Syne-1 P roper nuclear positioning relative to the cell body is important for many cellular processes during animal development. Mammalian skeletal muscle fibers are giant syncytial cells, each containing more than 100 nuclei. Each muscle fiber contains a neuromuscular junction (NMJ) that starts to form at approximately embryonic day 14 (E14) in mice (1). Following a multistep process involving the function of the agrin-MuSK-rapsynAChR pathway, the motor nerve terminal and the postsynaptic membrane become highly differentiated, ensuring reliable terminal transmission. The nuclei in the muscle fibers are positioned in a nonrandom manner, with 3-8 nuclei (synaptic nuclei) clustered beneath each NMJ and other nuclei (nonsynaptic nuclei) distributed evenly along the cell membrane (1, 2). How synaptic and nonsynaptic nuclei properly position themselves within each muscle fiber and the consequences of disrupting the nuclear positioning are attractive problems to investigate. Recent studies have indicated that KASH (Klarsicht/ANC-1/Syne homologue) domain-containing and SUN domain-containing proteins (KASH/SUN proteins) form a complex at the nuclear envelope (NE) for various cellular functions (3-6). Whereas a mammalian KASH protein has been found to have a critical function in myonuclear anchorage (7-9), the roles of SUN proteins in this process remain unclear.The SUN domain was first defined as a domain of shared homology between Sad1 in Schizosaccharomyces pombe and UNC-84 in Caenorhabditis elegans (10). SUN proteins are conserved inner nuclear membrane proteins containing at least 1 transmembrane domain and a C-terminal SUN...

Structural insights into SUN-KASH complexes across the nuclear envelope

“…By transducing mechanical force across the NE, the LINC complex has been implicated in many cellular events, such as nuclear migration and positioning [16,28]. Interestingly, the KASH protein Nesprin 2G has also been shown to positively regulate cell migration [25].…”

“…Both SUN1 and SUN2 are widely expressed KASH-binding partners and may have partially redundant functions. Double knockout of SUN1 and SUN2 is lethal in mice, similar to the disruption of KASH domain-containing proteins, highlighting the functional importance of the LINC complex [16].…”

The nuclear envelope at a glance

“…SUN-domain proteins, in turn, bind directly to lamins, forming a basic mechanical unit that spans the NE (Tzur et al, 2006). Sun1 and Sun2 have both overlapping and distinct roles (Lei et al, 2009;Olins et al, 2009). For example, Sun1 concentrates near NPCs and is required to space NPCs; in addition, Sun1 preferentially binds to the precursor form of lamin A, implying roles in lamin A maturation or assembly (Liu et al, 2007;Haque et al, 2010).…”

Muscle length and myonuclear position are independently regulated by distinct Dynein pathways

“…Moreover, mispositioned myonuclei, in conjunction with smaller myofibers, are a hallmark of many muscle disorders and have been used diagnostically for decades (Pierson et al, 2007;Romero, 2010). The mechanisms by which nuclei are moved throughout the muscle (Metzger et al, 2012) and anchored at the neuromuscular junction (NMJ) (Grady et al, 2005;Lei et al, 2009) are beginning to emerge; however, only a small number of factors necessary for these processes have been identified. Additionally, potential links between muscle size and myonuclear position have not been explored.…”