Defects in early secretory pathway transport machinery components and neurodevelopmental disorders

Abstract The early secretory pathway, provisionally comprising of vesicular traffic between the endoplasmic reticulum (ER) and the Golgi apparatus, occurs constitutively in mammalian cells. Critical for a constant supply of secretory and plasma membrane (PM) materials, the pathway is presumably essential for general cellular function and survival. Neurons exhibit a high intensity in membrane dynamics and protein/lipid trafficking, with differential and polarized trafficking towards the somatodendritic and axonal PM domains. Mutations in genes encoding early secretory pathway membrane trafficking machinery components are known to result in neurodevelopmental or neurological disorders with disease manifestation in early life. Here, such rare disorders associated with autosomal recessive mutations in coat proteins, membrane tethering complexes and membrane fusion machineries responsible for trafficking in the early secretory pathway are summarily discussed. These mutations affected genes encoding subunits of coat protein complex I and II, subunits of transport protein particle (TRAPP) complexes, members of the YIP1 domain family (YIPF) and a SNAP receptor (SNARE) family member. Why the ubiquitously present and constitutively acting early secretory pathway machinery components could specifically affect neurodevelopment is addressed, with the plausible underlying disease etiologies and neuropathological mechanisms resulting from these mutations explored.

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Yeast Golgi SNARE interactions are promiscuous

Journal of Cell Science ◽

10.1242/jcs.113.1.145 ◽

2000 ◽

Vol 113 (1) ◽

pp. 145-152 ◽

Cited By ~ 4

Author(s):

M.M. Tsui ◽

D.K. Banfield

Keyword(s):

Protein Interactions ◽

Membrane Trafficking ◽

Secretory Pathway ◽

Snare Proteins ◽

Current View ◽

Over Expression ◽

Vesicular Traffic ◽

Binding Data ◽

Genes Encoding ◽

Target Membrane

The transport of proteins between various compartments of the secretory pathway occurs by the budding of vesicles from one membrane and their fusion with another. A key event in this process is the selective recognition of the target membrane by the vesicle and the current view is that SNARE protein interactions likely play a central role in vesicle-target recognition and or membrane fusion. In yeast, only a single syntaxin (Sed5p) is required for Golgi transport and Sed5p is known to bind to at least 7 SNARE proteins. However, the number of Sed5p-containing SNARE complexes that exist in cells is not known. In this study we examined direct pair-wise interactions between full length soluble recombinant forms of SNAREs (Sed5p, Sft1p, Ykt6p, Vti1p, Gos1p, Sec22p, Bos1p, and Bet1p) involved in ER-Golgi and intra-Golgi membrane trafficking. In the binding assay that we describe here the majority of SNARE-binary interactions tested were positive, indicating that SNARE-SNARE interactions although promiscuous are not entirely non-selective. Interactions between a number of the genes encoding these SNAREs are consistent with our binding data and taken together our results suggest that functionally redundant Golgi SNARE-complexes exist in yeast. In particular, over-expression of Bet1p (a SNARE required for ER-Golgi and Golgi-ER traffic) and can bypass the requirement for the otherwise essential SNARE Sft1p (required for intra-Golgi traffic), suggesting that Bet1p either functions in a parallel pathway with Sft1p or can be incorporated into SNARE-complexes in place of Sftp1. None-the-less this result suggests that Bet1p can participate in two distinct trafficking steps, cycling between the ER and Golgi as well as in retrograde intra-Golgi traffic. In addition, suppressor genetics together with the analysis of the phenotypes of conditional mutations in Sft1p and Ykt6p, are consistent with a role for these SNAREs in more than one trafficking step. We propose that different combinations of SNAREs form complexes with Sed5p and are required for multiple steps in ER-Golgi and intra-Golgi vesicular traffic. And that the apparent promiscuity of SNARE-SNARE binding interactions, together with the requirement for some SNAREs in more than one trafficking step, supports the view that the specificity of vesicle fusion events cannot be explained solely on the basis of SNARE-SNARE interactions.

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Human ARF4 expression rescues sec7 mutant yeast cells.

Molecular and Cellular Biology ◽

10.1128/mcb.16.7.3275 ◽

1996 ◽

Vol 16 (7) ◽

pp. 3275-3284 ◽

Cited By ~ 23

Author(s):

S B Deitz ◽

C Wu ◽

S Silve ◽

K E Howell ◽

P Melançon ◽

...

Keyword(s):

Mammalian Cells ◽

Secretory Pathway ◽

Genetic Selection ◽

Yeast Cells ◽

Selection Strategy ◽

Restrictive Temperature ◽

Coat Proteins ◽

Yeast Saccharomyces Cerevisiae ◽

Interacting Protein ◽

Vesicular Traffic

Vesicle-mediated traffic between compartments of the yeast secretory pathway involves recruitment of multiple cytosolic proteins for budding, targeting, and membrane fusion events. The SEC7 gene product (Sec7p) is a constituent of coat structures on transport vesicles en route to the Golgi complex in the yeast Saccharomyces cerevisiae. To identify mammalian homologs of Sec7p and its interacting proteins, we used a genetic selection strategy in which a human HepG2 cDNA library was transformed into conditional-lethal yeast sec7 mutants. We isolated several clones capable of rescuing sec7 mutant growth at the restrictive temperature. The cDNA encoding the most effective suppressor was identified as human ADP ribosylation factor 4 (hARF4), a member of the GTPase family proposed to regulate recruitment of vesicle coat proteins in mammalian cells. Having identified a Sec7p-interacting protein rather than the mammalian Sec7p homolog, we provide evidence that hARF4 suppressed the sec7 mutation by restoring secretory pathway function. Shifting sec7 strains to the restrictive temperature results in the disappearance of the mutant Sec7p cytosolic pool without apparent changes in the membrane-associated fraction. The introduction of hARF4 to the cells maintained the balance between cytosolic and membrane-associated Sec7p pools. These results suggest a requirement for Sec7p cycling on and off of the membranes for cell growth and vesicular traffic. In addition, overexpression of the yeast GTPase-encoding genes ARF1 and ARF2, but not that of YPT1, suppressed the sec7 mutant growth phenotype in an allele-specific manner. This allele specificity indicates that individual ARFs are recruited to perform two different Sec7p-related functions in vesicle coat dynamics.

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Specific and Nonspecific Membrane-binding Determinants Cooperate in Targeting Phosphatidylinositol Transfer Protein β-Isoform to the MammalianTrans-Golgi Network

Molecular Biology of the Cell ◽

10.1091/mbc.e06-01-0089 ◽

2006 ◽

Vol 17 (6) ◽

pp. 2498-2512 ◽

Cited By ~ 33

Author(s):

Scott E. Phillips ◽

Kristina E. Ile ◽

Malika Boukhelifa ◽

Richard P.H. Huijbregts ◽

Vytas A. Bankaitis

Keyword(s):

Membrane Trafficking ◽

Mammalian Cells ◽

Secretory Pathway ◽

Membrane Binding ◽

Bound State ◽

Missense Mutations ◽

Cis Acting ◽

Domain Mapping ◽

Phosphatidylinositol Transfer ◽

Golgi Network

Phosphatidylinositol transfer proteins (PITPs) regulate the interface between lipid metabolism and specific steps in membrane trafficking through the secretory pathway in eukaryotes. Herein, we describe the cis-acting information that controls PITPβ localization in mammalian cells. We demonstrate PITPβ localizes predominantly to the trans-Golgi network (TGN) and that this localization is independent of the phospholipid-bound state of PITPβ. Domain mapping analyses show the targeting information within PITPβ consists of three short C-terminal specificity elements and a nonspecific membrane-binding element defined by a small motif consisting of adjacent tryptophan residues (the W202W203motif). Combination of the specificity elements with the W202W203motif is necessary and sufficient to generate an efficient TGN-targeting module. Finally, we demonstrate that PITPβ association with the TGN is tolerant to a range of missense mutations at residue serine 262, we describe the TGN localization of a novel PITPβ isoform with a naturally occurring S262Q polymorphism, and we find no other genetic or pharmacological evidence to support the concept that PITPβ localization to the TGN is obligately regulated by conventional protein kinase C (PKC) or the Golgi-localized PKC isoforms δ or ε. These latter findings are at odds with a previous report that conventional PKC-mediated phosphorylation of residue Ser262is required for PITPβ targeting to Golgi membranes.

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Distinct Roles for the Yeast Phosphatidylinositol 4-Kinases, Stt4p and Pik1p, in Secretion, Cell Growth, and Organelle Membrane Dynamics

Molecular Biology of the Cell ◽

10.1091/mbc.11.8.2673 ◽

2000 ◽

Vol 11 (8) ◽

pp. 2673-2689 ◽

Cited By ~ 254

Author(s):

Anjon Audhya ◽

Michelangelo Foti ◽

Scott D. Emr

Keyword(s):

Cell Growth ◽

Membrane Trafficking ◽

Secretory Pathway ◽

Membrane Dynamics ◽

Small Gtpase ◽

Effector Proteins ◽

Restrictive Temperature ◽

Yeast Saccharomyces Cerevisiae ◽

Cytoskeleton Organization ◽

Yeast Viability

The yeast Saccharomyces cerevisiae possesses two genes that encode phosphatidylinositol (PtdIns) 4-kinases,STT4 and PIK1. Both gene products phosphorylate PtdIns at the D-4 position of the inositol ring to generate PtdIns(4)P, which plays an essential role in yeast viability because deletion of either STT4 orPIK1 is lethal. Furthermore, although both enzymes have the same biochemical activity, increased expression of either kinase cannot compensate for the loss of the other, suggesting that these kinases regulate distinct intracellular functions, each of which is required for yeast cell growth. By the construction of temperature-conditional single and double mutants, we have found that Stt4p activity is required for the maintenance of vacuole morphology, cell wall integrity, and actin cytoskeleton organization. In contrast, Pik1p is essential for normal secretion, Golgi and vacuole membrane dynamics, and endocytosis. Strikingly,pik1tscells exhibit a rapid defect in secretion of Golgi-modified secretory pathway cargos, Hsp150p and invertase, whereas stt4tscells exhibit no detectable secretory defects. Both single mutants reduce PtdIns(4)P by ∼50%; however,stt4ts/pik1tsdouble mutant cells produce more than 10-fold less PtdIns(4)P as well as PtdIns(4,5)P2. The aberrant Golgi morphology found in pik1tsmutants is strikingly similar to that found in cells lacking the function of Arf1p, a small GTPase that is known to regulate multiple membrane trafficking events throughout the cell. Consistent with this observation, arf1 mutants exhibit reduced PtdIns(4)P levels. In contrast, diminished levels of PtdIns(4)P observed in stt4tscells at restrictive temperature result in a dramatic change in vacuole size compared with pik1tscells and persistent actin delocalization. Based on these results, we propose that Stt4p and Pik1p act as the major, if not the only, PtdIns 4-kinases in yeast and produce distinct pools of PtdIns(4)P and PtdIns(4,5)P2that act on different intracellular membranes to recruit or activate as yet uncharacterized effector proteins.

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Membrane Dynamics in the Early Secretory Pathway

Critical Reviews in Plant Sciences ◽

10.1080/07352680701495820 ◽

2007 ◽

Vol 26 (4) ◽

pp. 199-225 ◽

Cited By ~ 77

Author(s):

David G. Robinson ◽

Marie-Carmen Herranz ◽

Julia Bubeck ◽

Rainer Pepperkok ◽

Christophe Ritzenthaler

Keyword(s):

Secretory Pathway ◽

Membrane Dynamics ◽

Early Secretory Pathway

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DYRK3-Controlled Phase Separation Organizes the Early Secretory Pathway

10.1101/2020.02.10.941757 ◽

2020 ◽

Author(s):

Raffaella Gallo ◽

Arpan Rai ◽

Lucas Pelkmans

Keyword(s):

Mammalian Cells ◽

Secretory Pathway ◽

Dynamic Equilibrium ◽

Membrane Traffic ◽

Matrix Proteins ◽

Exit Site ◽

Early Secretory Pathway ◽

Physiological Processes ◽

Dual Specificity ◽

Peripheral Membrane Protein

SummaryThe dual-specificity kinase DYRK3 controls formation and dissolution of several intracellular condensates thereby regulating various cell physiological processes. Here we report that DYRK3 establishes a dynamic equilibrium between condensation and dissolution of proteins associated with membranous structures of the early secretory pathway to organize membrane traffic between the ER and the Golgi complex in mammalian cells. This depends on the peripheral membrane protein Sec16A, whose N-terminal disordered region forms DYRK3-controlled liquid-like condensates on the surface of the ER and co-phase separates with multiple ER exit site components and a subset of matrix proteins specifically associated with ERGIC and cis-Golgi. Our findings support a mechanism whereby multiple interacting and differentially regulated intracellular condensates create favorable environments for directional membrane traffic in eukaryotic cells.

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The role of microtubules in transport between the endoplasmic reticulum and Golgi apparatus in mammalian cells.

Biochemical Society Symposium ◽

10.1042/bss0720001 ◽

2005 ◽

Vol 72 ◽

pp. 1-13 ◽

Cited By ~ 24

Author(s):

Krysten J. Palmer ◽

Peter Watson ◽

David J. Stephens

Keyword(s):

Endoplasmic Reticulum ◽

Mammalian Cells ◽

Secretory Pathway ◽

Motor Proteins ◽

Microtubule Cytoskeleton ◽

Early Secretory Pathway ◽

Golgi Transport ◽

Intracellular Compartments ◽

Retrograde Traffic

The organization of intracellular compartments and the transfer of components between them are central to the correct functioning of mammalian cells. Proteins and lipids are transferred between compartments by the formation, movement and subsequent specific fusion of transport intermediates. These vesicles and membrane clusters must be coupled to the cytoskeleton and to motor proteins that drive motility. Anterograde ER (endoplasmic reticulum)-to-Golgi transport, and the converse step of retrograde traffic from the Golgi to the ER, are now known to involve coupling of membranes to the microtubule cytoskeleton. Here we shall discuss our current understanding of the mechanisms that link membrane traffic in the early secretory pathway to the microtubule cytoskeleton in mammalian cells. Recent data have also provided molecular detail of functional co-ordination of motor proteins to specify directionality, as well as mechanisms for regulating motor activity by protein phosphorylation.

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Involvement of Specific COPI Subunits in Protein Sorting from the Late Endosome to the Vacuole in Yeast

Molecular and Cellular Biology ◽

10.1128/mcb.00577-06 ◽

2006 ◽

Vol 27 (2) ◽

pp. 526-540 ◽

Cited By ~ 43

Author(s):

Galina Gabriely ◽

Rachel Kama ◽

Jeffrey E. Gerst

Keyword(s):

Mammalian Cells ◽

Secretory Pathway ◽

Protein Sorting ◽

Multivesicular Body ◽

Carboxypeptidase Y ◽

Direct Role ◽

Vacuolar Protein Sorting ◽

Early Secretory Pathway ◽

Physical Interactions ◽

Vacuolar Protein

ABSTRACT Although COPI function on the early secretory pathway in eukaryotes is well established, earlier studies also proposed a nonconventional role for this coat complex in endocytosis in mammalian cells. Here we present results that suggest an involvement for specific COPI subunits in the late steps of endosomal protein sorting in Saccharomyces cerevisiae. First, we found that carboxypeptidase Y (CPY) was partially missorted to the cell surface in certain mutants of the COPIB subcomplex (COPIb; Sec27, Sec28, and possibly Sec33), which indicates an impairment in endosomal transport. Second, integral membrane proteins destined for the vacuolar lumen (i.e., carboxypeptidase S [CPS1]; Fur4, Ste2, and Ste3) accumulated at an aberrant late endosomal compartment in these mutants. The observed phenotypes for COPIb mutants resemble those of class E vacuolar protein sorting (vps) mutants that are impaired in multivesicular body (MVB) protein sorting and biogenesis. Third, we observed physical interactions and colocalization between COPIb subunits and an MVB-associated protein, Vps27. Together, our findings suggest that certain COPI subunits could have a direct role in vacuolar protein sorting to the MVB compartment.

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