A CLN6-CLN8 complex recruits lysosomal enzymes at the ER for Golgi transfer

Bajaj, Lakshya; Ronza, Alberto di; Zheng, Ping; Eblimit, Aiden; Pal, Rituraj; Sharma, Jaiprakash; Roman, Dany; Collette, John R.; Sifers, Richard N.; Jung, Sung Yun; Chen, Rui; Schekman, Randy; Sardiello, Marco

doi:10.1101/773804

Cited by 7 publications

(13 citation statements)

References 74 publications

(75 reference statements)

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“…Given that the 239fsX human CLN6 mutant is prematurely terminated in the sixth transmembrane domain, a perturbation in CLN6 transmembrane domains is likely to affect the entire structure of CLN6 and in turn to impair its functionality. The third loop, predicted to face the ER lumen and also referred to as the second luminal loop, of CLN6 has been associated with ER-to-Golgi trafficking of a spectrum of lysosomal enzymes (Bajaj et al 2020). We previously demonstrated the requirement for the third loop in CLN6's anti-aggregate activity (Yamashita et al 2020).…”

Section: Removal Of Cln6's Luminal Tail Resulted In Insensitivity To the 132fsx Mutantmentioning

confidence: 99%

CLN6’s luminal tail-mediated functional interference between CLN6 mutants as a novel pathomechanism for the neuronal ceroid lipofuscinoses

et al. 2021

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CLN6 (Ceroid Lipofuscinosis, Neuronal, 6) is a 311-amino acid protein spanning the endoplasmic reticulum membrane. Mutations in CLN6 are linked to CLN6 disease, a hereditary neurodegenerative disorder categorized into the neuronal ceroid lipofuscinoses. CLN6 disease is an autosomal recessive disorder and individuals affected with this disease have two identical (homozygous) or two distinct (compound heterozygous) CLN6 mutant alleles. Little has been known about CLN6's physiological roles and the disease mechanism. We recently found that CLN6 prevents protein aggregate formation, pointing to impaired CLN6's anti-aggregate activity as a cause for the disease. To comprehensively understand the pathomechanism, overall anti-aggregate activity derived from two different CLN6 mutants needs to be investigated, considering patients compound heterozygous for CLN6 alleles. We focused on mutant combinations involving the S132CfsX18 (132fsX) prematurely terminated protein, produced from the most frequent mutation in CLN6. The 132fsX mutant nullified anti-aggregate activity of the P299L CLN6 missense mutant but not of wild-type CLN6. Wild-type CLN6's resistance to the 132fsX mutant was abolished by replacement of amino acids 297-301, including Pro297 and Pro299, with five alanine residues. Given that removal of CLN6's C-terminal fifteen amino acids 297-311 (luminal tail) did not affect the resistance, we suggested that CLN6's luminal tail, when unleashed from Pro297/299-mediated conformational constraints, is improperly positioned by the 132fsX mutant, thereby blocking the induction of antiaggregate activity. We here reveal a novel mechanism for dissipating CLN6 mutants' residual functions, providing an explanation for the compound heterozygosity-driven pathogenesis.

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Section: Removal Of Cln6's Luminal Tail Resulted In Insensitivity To the 132fsx Mutantmentioning

confidence: 99%

CLN6’s luminal tail-mediated functional interference between CLN6 mutants as a novel pathomechanism for the neuronal ceroid lipofuscinoses

et al. 2021

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“…Yet deletion of the p23/24 family of cargo receptors in yeast or pharmacological inhibition of their ability to interact with COPII in the cytosol in mammals leads to breakdown of the filtration platform at ERESs and the secretion of resident ER proteins (Belden and Barlowe, 2001;Ma et al, 2017). The coordinated selection of lysosomal enzymes for ER exit by the CLN6-CLN8 complex supports a role for the assembly of sorting platforms in ERESs, where CLN6 remains in the ER while CLN8, which binds COPII, is exported with its cargo enzymes (Bajaj et al, 2020).…”

Section: Exit From the Er: Sorting Platforms Gates And Filtersmentioning

confidence: 99%

A tango for coats and membranes: New insights into ER-to-Golgi traffic

Aridor

2022

Cell Reports

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The realization that the meticulous organization of cellular organelles is not required for the reconstitution of select intracellular traffic steps has revolutionized cell biology. It transformed the discipline from a morphological one into a molecular one. It helped in defining the activities of COPII and COPI vesicle coats in secretion. The work established the principles of the vesicular traffic model as envisioned by George Palade 50 years ago. However, in recent years, numerous advances in cellular and imaging technologies afforded an unprecedented molecular resolution that sheds new light on COPII activities and biosynthetic traffic between the ER and the Golgi. In the following review, I summarize this new information and attempt to provide a unified physical-molecular-morphological description of this traffic step. This information expands on the simplistic principles of vesicular traffic and provides novel frameworks to examine and explain physiological secretion. COAT-MEDIATED INTRACELLULAR TRAFFIC: CHALLENGING THE DOGMAIntracellular traffic is carried by vesicles-this dogma, pioneered by the work of George Palade, has guided the field for the past 50 years (Palade, 1975). Vesicular traffic has been documented across species throughout evolution and in the different cellular sorting sites during endo-and exocytosis (Mellman and Emr, 2013;Traub and Bonifacino, 2013). Vesicles bud from donor membranes, travel in space, tether, and fuse with acceptor membranes to deliver cargoes (Figure 1). Vesicles are formed by the activity of coats, cytosolic protein complexes that are recruited to donor membranes to bud vesicles. Recruitment is mediated by small guanosine triphosphatases (GTPases) (secretion) or through activation by facilitators that drive conformational change (internalization; Kirchhausen et al., 2014;Miller and Schekman, 2013). Once recruited, coats interact with membranes and cargoes. Through such interactions, the coats sort and collect cargoes, tethers, and fusion machinery into membrane domains that are molded into buds and vesicles with help from coat polymerization. Once vesicles bud and uncoat, tether and fusion activities will deliver cargo to the acceptor membrane. This dogma was established from robust combinations of data using genetic, cellular, and biochemical approaches in cells and cell-free reconstitution systems (Kirchhausen et al., 2014;Miller and Schekman, 2013). Yet recent work focusing on one of the key sorting sites in cells at ER exit sites that were monitored in an unprecedented resolution suggest otherwise (Shomron et al., 2021;Weigel et al., 2021). Instead of vesicles, connectivity between donor and acceptor membranes may drive traffic (Figure 1C), and ER exit site components, including TANGO1, direct such connectivity (Raote et al., 2021; Figure 2). Instead of coat-mediated vesicle formation, coat-assisted filters are proposed, sorting cargoes at an ER exit site gate. We may even have two coats, not one, contributing to the formation of one single tubular transport carri...

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“…CRMP2 and KLC4 have well established functions in axonal transport, but roles for CLN6 in this process have not been described. Since CLN6 is a resident ER transmembrane protein involved in ER to Golgi vesicle transport of lysosomal enzymes, we asked whether the CCK complex could be regulating the axonal transport of ER-derived vesicles [9,27,31,32]. Mouse cortical neurons were cultured and labeled for ER-derived vesicles in live cells with ER-tracker.…”

Section: Cln6 Interacts With Crmp2 and Klc4 To Regulate Axonal Transport Of Er-derived Vesiclesmentioning

confidence: 99%

A CLN6-CRMP2-KLC4 complex regulates anterograde ER-derived vesicle trafficking in cortical neurites

Koh

Cain

Magee

et al. 2021

Preprint

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As neurons establish extensive connections throughout the central nervous system, the transport of cargo along the microtubule network of the axon is crucial for differentiation and homeostasis. Specifically, building blocks such as membrane and cytoskeletal components, organelles, transmembrane receptors, adhesion molecules, and peptide neurotransmitters all require proper transport to the presynaptic compartment. Here, we identify a novel complex regulating vesicular endoplasmic reticulum transport in neurites, composed of CLN6: an ER-associated protein of relatively unknown function implicated in CLN6-Batten disease; CRMP2: a tubulin binding protein important in regulating neurite microtubule dynamics; and KLC4: a classic transport motor protein. We show that this 'CCK' complex allows ER-derived vesicles to migrate to the distal end of the axon, aiding in proper neurite outgrowth and arborization. In the absence of CLN6, the CCK complex does not function effectively, leading to reduced vesicular transport, stunted neurite outgrowth, and deficits in CRMP2 binding to other protein partners. Treatment with a CRMP2 modulating compound, lanthionine ketimine ester, partially restores these deficits in CLN6-deficient mouse neurons, indicating that stabilization of CRMP2 interacting partners may prove beneficial in lieu of complete restoration of the CCK complex. Taken together, these findings reveal a novel mechanism of ER-derived vesicle transport in the axon and provide new insights into therapeutic targets for neurodegenerative disease.

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A CLN6-CLN8 complex recruits lysosomal enzymes at the ER for Golgi transfer

Cited by 7 publications

References 74 publications

CLN6’s luminal tail-mediated functional interference between CLN6 mutants as a novel pathomechanism for the neuronal ceroid lipofuscinoses

CLN6’s luminal tail-mediated functional interference between CLN6 mutants as a novel pathomechanism for the neuronal ceroid lipofuscinoses

A tango for coats and membranes: New insights into ER-to-Golgi traffic

A CLN6-CRMP2-KLC4 complex regulates anterograde ER-derived vesicle trafficking in cortical neurites

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