DOC_MIT_MIM_1

Cells in eukaryotes have complex endomembrane systems and sorting events that require membrane remodelling complexes. ESCRT (endosomal sorting complex required for transport) machinery performs membrane remodelling, sorting and scission events. This machinery consists of a series of protein complexes, including ESCRT-0, -I, -II, -III, and an AAA ATPase VPS4. VPS4 performs the sequential recruitment of ESCRT-III layers, turnover, and disassembly of subunits in an ATP-dependent manner (Pfitzner,2020; Mierzwa,2017). The ESCRT-III complex is responsible for the Intraluminal vesicle (ILVs) formation during the multivesicular body (MVB) formation (Katzmann,2002), cytokinetic abscission during the last stages of cytokinesis (Carlton,2007), reformation of the nuclear envelope and sealing (Vietri,2015), budding of virions (including HIV-1) in eukaryotes (Morita,2004), neuron pruning (Loncle,2015) and cortical constriction (Elia,2011). The metazoan ESCRT-III subunits (CHMP1-7, IST1) polymerise into spiral filaments to constrict the vesicle neck region during the egress of viruses via the secretory pathway or during the budding of intraluminal vesicles. This process is known as reverse topology membrane fission, which results in the budding away of the vesicle from the cytosolic surface of the membrane (Pfitzner,2020; Schoneberg,2017).

A number of ESCRT complex proteins (VPS4, Vta1, LIP5, Spartin, ULK3, AMSH, UBPY, MITD1, and Spastin) contain the MIT (microtubule interacting and trafficking) domain. The MIT domain is a three-helix bundle that interacts with proteins through different surface grooves formed by its three helices. MIMs (MIT Interacting Motifs) present in ESCRT-III subunits (CHMP1A, CHMP1B, CHMP2A, IST1, VPS2, DID2) dock to the MIT domain. Five types of MIMs (MIM1-5) have been reported to interact with different sites/surface grooves on the MIT domain. MIM1 is an α-helical motif, and its central Leu residues form hydrophobic contacts in the region between the α2 and α3 helix of the MIT domain. Polar residues flank the MIM1 motif core and engage with the MIT domain via H-bonds and salt bridge interactions (Guo,2015; Obita,2007; Stuchell-Brereton,2007). MIM2 contains at least two essential prolines and binds as a random coil between α1 and α3 of the MIT domain (Kieffer,2008; Vild,2014). MIM3 interacts as a helical structure in the same region, and its C-terminus is similar to MIM1 as observed in the CHMP1B - Spastin complex (Yang,2008). MIM4 also adopts a helical conformation and binds in the groove generated by helices 3 and 4/5 of the AMSH MIT domain (Solomons,2011). MIM5 forms two helices and interacts with MIT domain helices 1-3. A representative MIM5 interaction is observed in VPS60 (128–186) - Vta1 NTD (N-terminal Domain) complex (Yang,2012).

ESCRT–III protein IST1 (Increased sodium tolerance-1; P53990) is required for cytokinetic abscission and contains MIM1 and MIM2 docking motifs. Replacing key interacting residues (L375A/K376A) of IST1 MIM1 resulted in defective multinucleated phenotypes, indicating the importance of this region (Agromayor,2009; Bajorek,2009). Further, MIM1 and MIM2 motifs are employed by IST1 to bind MIT domain-containing protein kinase ULK3 (Unc-51-like kinase 3). This interaction results in IST1 phosphorylation by ULK3, thereby delaying cytokinetic abscission in response to lagging chromosomes in the mid-body during cell division (Caballe,2015). Interestingly, IST1 can bind in both MIM1, and MIM3 modes as the bulky side-chain of Phe357 provides sufficient binding energy while interacting with the MIT domain of VPS4 (4U7Y) and Spartin (4U7I), respectively (Guo,2015). Point mutations in the key MIT interacting residues of the IST1 MIM1 (L353A, R357E, L360A, L360A/K361A; P53990) abrogates the interaction with the MIT domain of Spartin, LIP5, VPS4B, VPS4, MITD1, UBPY (Guo,2015; Bajorek,2009; Agromayor,2009).

MIM1-MIT binding between MITD1 (MIT-domain containing protein 1) (Q8WV92) and ESCRT-III subunits (IST1, CHMP1A, CHMP1B, CHMP2A) allows the recruitment of MITD1 during cell division (Hadders,2012). MIM1,2-MIT interaction regulates both the recruitment and functioning of VPS4 ATPase (Han,2015; Shestakova,2010). Substituting the critical Leu and Arg residues (L221D, R224D, L225D, L228D) in the VPS2 MIM1 motif impedes its binding with VPS4 (Obita,2007).

MIM1 forms an amphipathic helix where one side of the alpha helix makes hydrophobic interactions with the MIT domain, whereas the other side engages in hydrophilic contacts. The former interactions happen via Leu residues in the MIM1 core region, whereas the latter interactions happen through aspartic acid, glutamic acid at N-terminus and Arg, Lys at the C terminus (2V6X; 2JQ9; 2JQK; 4WZX; 4U7E; 4U7I; 4U7Y).