Structural role of essential light chains in the apicomplexan glideosome

Abstract

28Apicomplexan parasites, such as Plasmodium falciparum and Toxoplasma gondii, traverse 29 the host tissues and invade the host cells exhibiting a specific type of motility called gliding. 30The molecular mechanism of gliding lies in the actin-myosin motor localized to the 31 intermembrane space between the plasma membrane and inner membrane complex (IMC) of 32 the parasites. Myosin A (MyoA) is a part of the glideosome, a large multi-protein complex, 33 which is anchored in the outer membrane of the IMC. MyoA is bound to the proximal essential 34 light chain (ELC) and distal myosin light chain (MLC1), which further interact with the 35 glideosome associated proteins GAP40, GAP45 and GAP50. Whereas structures of several 36 individual glideosome components and small dimeric complexes have been solved, structural 37 information concerning the interaction of larger glideosome subunits and their role in 38 glideosome function still remains to be elucidated. Here, we present structures of a T. gondii 39 trimeric glideosome sub complex composed of a myosin A light chain domain with bound 40 MLC1 and TgELC1 or TgELC2. Regardless of the differences between the secondary 41 structure content observed for free P. falciparum PfELC and T. gondii TgELC1 or TgELC2, 42 the proteins interact with a conserved region of TgMyoA to form structurally conserved 43 complexes. Upon interaction, the essential light chains undergo contraction and induce 44 α-helical structure in the myosin A C-terminus, stiffening the myosin lever arm. The complex 45 formation is further stabilized through binding of a single calcium ion to T. gondii ELCs. Our 46 work provides an important step towards the structural understanding of the entire glideosome 47 and uncovering the role of its members in parasite motility and invasion. 48 49 50 3 51 Author summary 52 Apicomplexans, such as Toxoplasma gondii or the malaria agent Plasmodium falciparum, are 53 small unicellular parasites that cause serious diseases in humans and other animals. These 54 parasites move and infect the host cells by a unique type of motility called gliding. Gliding is 55 empowered by an actin-myosin molecular motor located at the periphery of the parasites. 56 Myosin interacts with additional proteins such as essential light chains to form the glideosome, 57 a large protein assembly that anchors myosin in the inner membrane complex. Unfortunately, 58 our understanding of the glideosome is insufficient because we lack the necessary structural 59 information. Here we describe the first structures of trimeric glideosome sub complexes of T. 60 gondii myosin A bound to two different light chain combinations, which show that T. gondii and 61 P. falciparum form structurally conserved complexes. With an additional calcium-free complex 62 structure, we demonstrate that calcium binding does not change the formation of the 63 complexes, although it provides them with substantial stability. With additional data, we 64 propose that the role of the essential light chains is to enhance myosin perfor...

Blocking palmitoylation ofToxoplasma gondiimyosin light chain 1 disrupts glideosome composition but has little impact on parasite motility

“…In the linear motor model of motility, TgMLC1 plays two key roles in force generation: it stabilizes the TgMyoA lever arm (10,13,20), and it serves as a physical linker connecting the TgMyoA motor to TgGAP45 and the IMC (22,23). We show here that TgMLC1 is dually palmitoylated, on C8 and C11 (Fig.…”

“…Alternatively, by inserting into the IMC membrane, the acyl chains could position the relevant N-terminal residues of TgMLC1 favorably for interaction with the C-terminus of TgGAP45, which is itself attached to the IMC membrane via palmitoylation. It is also possible that the acyl chains on TgMLC1 and TgGAP45 themselves interact within the plane of the membrane (20). In any of these cases, blocking TgMLC1 palmitoylation would be expected to inhibit TgMLC1-TgGAP45 interaction, as observed.…”

“…Direct protein-protein interaction may be involved, as the mutation of two other sets of conserved N-terminal amino acids in TgMLC1 (D26E28 and P36GF38) also interfere with binding to TgGAP45 (23). The presence of the palmitates on C8 and C11 of TgMLC1 might help the interacting N-terminal residues of TgMLC1 transition from a disordered state (13,20) into a binding-competent configuration. Alternatively, by inserting into the IMC membrane, the acyl chains could position the relevant N-terminal residues of TgMLC1 favorably for interaction with the C-terminus of TgGAP45, which is itself attached to the IMC membrane via palmitoylation.…”

“…TgMLC1 is thought to play two key roles within the T. gondii glideosome. First, TgMLC1 binds to the C-terminal tail of TgMyoA to reinforce the motor's lever arm (10,13,20). The lever arm amplifies small motions at the myosin active site into larger movements that are capable of displacing actin filaments (10,21).…”

Structure of Full Length Plasmodium Myosin A and its light chain PfELC, dual targets against malaria parasite pathogenesis

“…Atomic structures of the PfMyoA lever arm indicate how degenerate IQ motifs of the HC are recognized by PfELC and MTIP, as well as how unusual apolar interactions occur between PfELC and the Converter. These unusual features could not have been predicted from recently solved TgMyoA structures of the lever arm bound to TgELC or TgELC2 31 , because TgELC is a poor model for PfELC considering the low sequence identity (20 and 21% between PfELC and TgELC and TgELC2, respectively). Interestingly, TgELC1 and TgELC2 are homologous to classical ELCs and do not display the specific inter-lobe linker nor the atypical α5* and α5'* helices found in PfELC, which are also part of the interface with the Converter.…”