Loss of Arabidopsis p24 function affects ERD2 traffic and Golgi structure and activates the unfolded protein response
The p24 family of proteins (also known as the TMED family) are key regulators of protein trafficking along the secretory pathway, but very little is known about their functions in plants. A quadruple loss-of-function mutant affecting the p24 genes from the δ-1 subclass of the p24δ subfamily (p24δ3δ4δ5δ6) showed alterations in the Golgi, suggesting that these p24 proteins play a role in the organization of the compartments of the early secretory pathway in Arabidopsis Loss of p24δ-1 proteins also induced the accumulation of the K/HDEL receptor ERD2a (ER lumen protein-retaining receptor A) at the Golgi and increased secretion of BiP family proteins, ER chaperones containing an HDEL signal, pr…
alpha 2-COP is involved in early secretory traffic in Arabidopsis and is required for plant growth
[EN] COP (coat protein) I-coated vesicles mediate intra-Golgi transport and retrograde transport from the Golgi to the endoplasmic reticulum. These vesicles form through the action of the small GTPase ADP-ribosylation factor 1 (ARF1) and the COPI heptameric protein complex (coatomer), which consists of seven subunits (alpha-, beta-, beta' -, gamma-, delta-, epsilon- and xi-COP). In contrast to mammals and yeast, several isoforms for coatomer subunits, with the exception of gamma and delta, have been identified in Arabidopsis. To understand the role of COPI proteins in plant biology, we have identified and characterized a loss-of-function mutant of alpha 2-COP, an Arabidopsis alpha-COP isofo…
p24 Family Proteins Are Involved in Transport to the Plasma Membrane of GPI-Anchored Proteins in Plants
p24 proteins are a family of type-I membrane proteins that cycle between the endoplasmic reticulum (ER) and the Golgi apparatus via Coat Protein I (COPI)- and COPII-coated vesicles. These proteins have been proposed to function as cargo receptors, but the identity of putative cargos in plants is still elusive. We previously generated an Arabidopsis (Arabidopsis thaliana) quadruple loss-of-function mutant affecting p24 genes from the δ-1 subclass of the p24 delta subfamily (p24δ3δ4δ5δ6 mutant). This mutant also had reduced protein levels of other p24 family proteins and was found to be sensitive to salt stress. Here, we used this mutant to test the possible involvement of p24 proteins in the…
AtPGAP1 functions as a GPI inositol-deacylase required for efficient transport of GPI-anchored proteins
Abstract Glycosylphosphatidylinositol (GPI)-anchored proteins (GPI-APs) play an important role in a variety of plant biological processes including growth, stress response, morphogenesis, signaling, and cell wall biosynthesis. The GPI anchor contains a lipid-linked glycan backbone that is synthesized in the endoplasmic reticulum (ER) where it is subsequently transferred to the C-terminus of proteins containing a GPI signal peptide by a GPI transamidase. Once the GPI anchor is attached to the protein, the glycan and lipid moieties are remodeled. In mammals and yeast, this remodeling is required for GPI-APs to be included in Coat Protein II-coated vesicles for their ER export and subsequent t…
Transient Transformation of A. thaliana Seedlings by Vacuum Infiltration
Transient expression in Arabidopsis thaliana seedlings allows fast expression of fluorescent markers for different subcellular compartments. This protocol describes a transient transformation assay with five-day-old seedlings using Agrobacterium tumefaciens-mediated vacuum infiltration. Three days after infiltration of the Agrobacterium containing an expression vector for a fluorescent marker of interest, cotyledon cells expressing the fluorescent protein can be imaged in a confocal microscope. This assay allows high-throughput screening of new constructs and the study of the localization of a large number of subcellular markers in Arabidopsis seedlings including wild-type, stable over-expr…
N-Linked Glycosylation of the p24 Family Protein p24δ5 Modulates Retrograde Golgi-to-ER Transport of K/HDEL Ligands in Arabidopsis
Abstract The K/HDEL receptor ERD2 mediates the transport of soluble endoplasmic reticulum (ER)-resident proteins containing a C-terminal K/HDEL signal from the Golgi apparatus back to the ER via COPI (COat Protein I)-coated vesicles. Sorting of ERD2 within COPI vesicles is facilitated by p24 proteins. In Arabidopsis , p24δ5 has been shown to interact directly with ERD2 via its luminal GOLD (GOLgi Dynamics) domain and with COPI proteins via its cytoplasmic C-terminal tail at the acidic pH of the Golgi apparatus. Several members of the p24 family in mammals and yeast have been shown to be glycosylated, but whether Arabidopsis p24 proteins are glycosylated and the role of the sugar moiety in p…
Loss of
The early secretory pathway involves bidirectional transport between the endoplasmic reticulum (ER) and the Golgi apparatus and is mediated by coat protein complex I (COPI)-coated and coat protein complex II (COPII)-coated vesicles. COPII vesicles are involved in ER to Golgi transport meanwhile COPI vesicles mediate intra-Golgi transport and retrograde transport from the Golgi apparatus to the ER. The key component of COPI vesicles is the coatomer complex, that is composed of seven subunits (α/β/β'/γ/δ/ε/ζ). In Arabidopsis two genes coding for the β-COP subunit have been identified, which are the result of recent tandem duplication. Here we have used a loss-of-function approach to study the…
ß-COP mutants show specific high sensitivity to chloride ions.
Coat Protein I (COPI) consists of a complex (coatomer) formed by seven subunits (α-, β-, β’-, γ-, δ-, ε-, and ζ-COP) that is recruited to Golgi membranes to form vesicles that shuttle from the Golgi apparatus to the ER and between Golgi stacks. Recently, it has been described that loss of function mutants of the two Arabidopsis β-COP genes, β1-COP and β2-COP, showed increased sensitivity to salt stress (NaCl). Using a mixture of either Na(+) or Cl(−) salts, we have now found that β-COP mutants are specifically and highly sensitive to chloride ions.
Function of AtPGAP1 in GPI anchor lipid remodeling and transport to the cell surface of GPI-anchored proteins
ABSTRACTGPI-anchored proteins (GPI-APs) play an important role in a variety of plant biological processes including growth, stress response, morphogenesis, signalling and cell wall biosynthesis. The GPI-anchor contains a lipid-linked glycan backbone that is synthesized in the endoplasmic reticulum (ER) where it is subsequently transferred to the C-terminus of proteins containing a GPI signal peptide by a GPI transamidase. Once the GPI anchor is attached to the protein, the glycan and lipid moieties are remodelled. In mammals and yeast, this remodelling is required for GPI-APs to be included in Coat Protein II (COPII) coated vesicles for their ER export and subsequent transport to the cell s…