A highly sensitive Golgi-targeted fluorescent probe for the simultaneous detection of malondialdehyde and formaldehyde in living systems and foods.

Malondialdehyde (MDA) and formaldehyde (FA) are highly active carbonyl substances widely present in both biological and abiotic systems. The detection of MDA and FA is of great significance for disease diagnosis and food safety monitoring. However, due to the similarity in structural properties between MDA and FA, very few probes for synergistically detecting MDA and FA were reported. In addition, functional abnormalities in the Golgi apparatus are closely related to MDA and FA, but currently there are no fluorescent probes that can detect MDA and FA in the Golgi apparatus. Therefore, we constructed a simple Golgi-targetable fluorescent probe GHA based on hydrazine moiety as the recognition site to produce a pyrazole structure after reaction with MDA and to generate a CN double bond after reaction with FA, allowing MDA and FA to be distinguished due to different emission wavelengths during the recognition process. The probe GHA has good specificity and sensitivity. Under the excitation of 350 nm, the blue fluorescence was significantly enhanced at 424 nm when the probe reacted with MDA, and the detection limit was 71 nM. At the same time, under the same excitation of 350 nm, the reaction with FA showed a significant enhancement of green fluorescence at 520 nm, with a detection limit of 12 nM for FA. And the simultaneous and high-resolution imaging of MDA and FA in the Golgi apparatus of cells was achieved. In addition, the applications of the probe GHA in food demonstrated it can provide a powerful method for food safety monitoring. In summary, this study offers a promising tool for the synergistic identification and determination of MDA and FA in the biosystem and food, facilitating the revelation of their detailed functions in Golgi apparatus and the monitoring of food safety.

A golgi targeting viscosity rotor for cancer diagnosis in living cells and tissues.

Unveiling the intricate relationship between cancer and Golgi viscosity remains an arduous endeavor, primarily due to the lack of Golgi-specific fluorescent probes tailored for viscosity measurement. Considering this formidable obstacle, we have triumphed over the challenge by devising a bespoke Golgi-specific viscosity probe, aptly named GOL-V. This ingenious innovation comprises the viscosity rotor BODIPY intricately tethered to the Golgi-targeting moiety benzsulfamide. GOL-V exhibits remarkable sensitivity to fluctuations in viscosity, the fluorescence intensity of GOL-V increased 114-fold when the viscosity value was increased from 2.63 to 937.28 cP. Owing to its remarkable capacity to suppress the TICT state under conditions of heightened viscosity. Moreover, its efficacy in sensitively monitoring Golgi viscosity alterations within living cells is also very significant. Astonishingly, our endeavors have culminated in not only the visualization of Golgi viscosity at the cellular and tissue levels but also in the clinical tissue samples procured from cancer patients. Harnessing the prowess of GOL-V, we have successfully demonstrated that Golgi viscosity could serve as a discerning marker for detecting the presence of cancer. The convergence of these exceptional attributes firmly establishes GOL-V as an immensely potent instrument, holding immense potential in the realm of cancer diagnosis.

Proteomic dissection of large extracellular vesicle surfaceome unravels interactive surface platform.

The extracellular vesicle (EV) surface proteome (surfaceome) acts as a fundamental signalling gateway by bridging intra- and extracellular signalling networks, dictates EVs' capacity to communicate and interact with their environment, and is a source of potential disease biomarkers and therapeutic targets. However, our understanding of surface protein composition of large EVs (L-EVs, 100-800 nm, mean 310 nm, ATP5F1A, ATP5F1B, DHX9, GOT2, HSPA5, HSPD1, MDH2, STOML2), a major EV-subtype that are distinct from small EVs (S-EVs, 30-150 nm, mean 110 nm, CD44, CD63, CD81, CD82, CD9, PDCD6IP, SDCBP, TSG101) remains limited. Using a membrane impermeant derivative of biotin to capture surface proteins coupled to mass spectrometry analysis, we show that out of 4143 proteins identified in density-gradient purified L-EVs (1.07-1.11 g/mL, from multiple cancer cell lines), 961 proteins are surface accessible. The surface molecular diversity of L-EVs include (i) bona fide plasma membrane anchored proteins (cluster of differentiation, transporters, receptors and GPI anchored proteins implicated in cell-cell and cell-ECM interactions); and (ii) membrane surface-associated proteins (that are released by divalent ion chelator EDTA) implicated in actin cytoskeleton regulation, junction organization, glycolysis and platelet activation. Ligand-receptor analysis of L-EV surfaceome (e.g., ITGAV/ITGB1) uncovered interactome spanning 172 experimentally verified cognate binding partners (e.g., ANGPTL3, PLG, and VTN) with highest tissue enrichment for liver. Assessment of biotin inaccessible L-EV proteome revealed enrichment for proteins belonging to COPI/II-coated ER/Golgi-derived vesicles and mitochondria. Additionally, despite common surface proteins identified in L-EVs and S-EVs, our data reveals surfaceome heterogeneity between the two EV-subtype. Collectively, our study provides critical insights into diverse proteins operating at the interactive platform of L-EVs and molecular leads for future studies seeking to decipher L-EV heterogeneity and function.

A combinatorial method to visualize the neuronal network in the mouse spinal cord: combination of a modified Golgi-Cox method and synchrotron radiation micro-computed tomography.

Exploring the three-dimensional (3D) morphology of neurons is essential to understanding spinal cord function and associated diseases comprehensively. However, 3D imaging of the neuronal network in the broad region of the spinal cord at cellular resolution remains a challenge in the field of neuroscience. In this study, to obtain high-resolution 3D imaging of a detailed neuronal network in the mass of the spinal cord, the combination of synchrotron radiation micro-computed tomography (SRµCT) and the Golgi-cox staining were used. We optimized the Golgi-Cox method (GCM) and developed a modified GCM (M-GCM), which improved background staining, reduced the number of artefacts, and diminished the impact of incomplete vasculature compared to the current GCM. Moreover, we achieved high-resolution 3D imaging of the detailed neuronal network in the spinal cord through the combination of SRµCT and M-GCM. Our results showed that the M-GCM increased the contrast between the neuronal structure and its surrounding extracellular matrix. Compared to the GCM, the M-GCM also diminished the impact of the artefacts and incomplete vasculature on the 3D image. Additionally, the 3D neuronal architecture was successfully quantified using a combination of SRµCT and M-GCM. The SRµCT was shown to be a valuable non-destructive tool for 3D visualization of the neuronal network in the broad 3D region of the spinal cord. Such a combinatorial method will, therefore, transform the presentation of Golgi staining from 2 to 3D, providing significant improvements in the 3D rendering of the neuronal network.

Aminoglycerophospholipid flipping and P4-ATPases in Toxoplasma gondii.

Lipid flipping in the membrane bilayers is a widespread eukaryotic phenomenon that is catalyzed by assorted P4-ATPases. Its occurrence, mechanism, and importance in apicomplexan parasites have remained elusive, however. Here we show that Toxoplasma gondii, an obligate intracellular parasite with high clinical relevance, can salvage phosphatidylserine (PtdSer) and phosphatidylethanolamine (PtdEtn) but not phosphatidylcholine (PtdCho) probes from its milieu. Consistently, the drug analogs of PtdCho are broadly ineffective in the parasite culture. NBD-PtdSer imported to the parasite interior is decarboxylated to NBD-PtdEtn, while the latter is not methylated to yield PtdCho, which confirms the expression of PtdSer decarboxylase but a lack of PtdEtn methyltransferase activity and suggests a role of exogenous lipids in membrane biogenesis of T. gondii. Flow cytometric quantitation of NBD-probes endorsed the selectivity of phospholipid transport and revealed a dependence of the process on energy and protein. Accordingly, our further work identified five P4-ATPases (TgP4-ATPase1-5), all of which harbor the signature residues and motifs required for phospholipid flipping. Of the four proteins expressed during the lytic cycle, TgP4-ATPase1 is present in the apical plasmalemma; TgP4-ATPase3 resides in the Golgi network along with its noncatalytic partner Ligand Effector Module 3 (TgLem3), whereas TgP4-ATPase2 and TgP4-ATPase5 localize in the plasmalemma as well as endo/cytomembranes. Last but not least, auxin-induced degradation of TgP4-ATPase1-3 impaired the parasite growth in human host cells, disclosing their crucial roles during acute infection. In conclusion, we show selective translocation of PtdEtn and PtdSer at the parasite surface and provide the underlying mechanistic and physiological insights in a model eukaryotic pathogen.

A Golgi-Targeting and Dual-Color "Turn-On" Probe for Spatially Precise Imaging of Furin.

Development of fluorescence probes for highly accurate detection of cancer-related enzyme activity is important in early cancer diagnosis. Herein, we report a Golgi-targeting and dual-color "Turn-On" probe Q-RVRR-DCM for imaging furin with high spatial precision. By integrating the principles of Förster resonance energy transfer and intramolecular charge transfer, the probe was designed to be non-fluorescent. Upon furin cleavage, Q-RVRR-DCM was converted into Q-RVRR and DCM-NH2, turning the dual fluorescence color "On" at 420 and 640 nm without spectral cross-talk. In furin-overexpressing HCT116 cells, Q-RVRR-DCM showed not only furin-specific, dual-color "Turn-On" fluorescence but also superior colocalization with a Golgi tracker than the single-color "Turn-On" probe RVRR-DCM. We envision that, with the excellent properties of Golgi-targeting and dual fluorescence color "Turn-On", our furin probe Q-RVRR-DCM could be applied for accurate early diagnosis of cancer in the near future.

Living on the edge: the role of Atgolgin-84A at the plant ER-Golgi interface.

The plant Golgi apparatus is responsible for the processing of proteins received from the endoplasmic reticulum (ER) and their distribution to multiple destinations within the cell. Golgi matrix components, such as golgins, have been identified and suggested to function as putative tethering factors to mediate the physical connections between Golgi bodies and the ER network. Golgins are proteins anchored to the Golgi membrane by the C-terminus either through transmembrane domains or interaction with small regulatory GTPases. The golgin N-terminus contains long coiled-coil domains, which consist of a number of α-helices wrapped around each other to form a structure similar to a rope being made from several strands, reaching into the cytoplasm. In animal cells, golgins are also implicated in specific recognition of cargo at the Golgi.Here, we investigate the plant golgin Atgolgin-84A for its subcellular localization and potential role as a tethering factor at the ER-Golgi interface. For this, fluorescent fusions of Atgolgin-84A and an Atgolgin-84A truncation lacking the coiled-coil domains (Atgolgin-84AΔ1-557) were transiently expressed in tobacco leaf epidermal cells and imaged using high-resolution confocal microscopy. We show that Atgolgin-84A localizes to a pre-cis-Golgi compartment that is also labelled by one of the COPII proteins as well as by the tether protein AtCASP. Upon overexpression of Atgolgin-84A or its deletion mutant, transport between the ER and Golgi bodies is impaired and cargo proteins are redirected to the vacuole. LAY DESCRIPTION: The Golgi apparatus is a specialised compartment found in mammalian and plant cells. It is the post office of the cell and packages proteins into small membrane boxes for transport to their destination in the cell. The plant Golgi apparatus consist of many separate Golgi bodies and is responsible for the processing of proteins received from the endoplasmic reticulum (ER) and their distribution to multiple destinations within the cell. Specialised proteins called golgins have been suggested to tether Golgi bodies and the ER. Here we investigate the plant golgin Atgolgin-84A for its exact within the Golgi body and its potential role as a tethering factor at the ER-Golgi interface. For this, we have fused Atgolgin-84A with a fluorescent protein from jellyfish and we are producing this combination in tobacco leaf cells. This allows us to see the protein using laser microscopy. We show that Atgolgin-84A localises to a compartment between the ER and Golgi that is also labelled by the tether protein AtCASP. When Atgolgin-84A is produced in high amounts in the cell, transport between the ER and Golgi bodies is inhibited and proteins are redirected to the vacuole.

IGF-1 receptor activity in the Golgi of migratory cancer cells depends on adhesion-dependent phosphorylation of Tyr1250 and Tyr1251.

Although insulin-like growth factor 1 (IGF-1) signaling promotes tumor growth and cancer progression, therapies that target the IGF-1 receptor (IGF-1R) have shown poor clinical efficacy. To address IGF-1R activity in cancer cells and how it differs from that of the closely related insulin receptor (IR), we focused on two tyrosines in the IGF-1R C-terminal tail that are not present in the IR and are essential for IGF-1-mediated cancer cell survival, migration, and tumorigenic growth. We found that Tyr1250 and Tyr1251 (Tyr1250/1251) were autophosphorylated in a cell adhesion-dependent manner. To investigate the consequences of this phosphorylation, we generated phosphomimetic Y1250E/Y1251E (EE) and nonphosphorylatable Y1250F/Y1251F (FF) mutant forms of IGF-1R. Although fully competent in kinase activity and signaling, the EE mutant was more rapidly internalized and degraded than either the wild-type or FF receptor. IGF-1 promoted the accumulation of wild-type and EE IGF-1R within the Golgi apparatus, whereas the FF mutant remained at the plasma membrane. Golgi-associated IGF-1R signaling was a feature of migratory cancer cells, and Golgi disruption impaired IGF-1-induced signaling and cell migration. Upon the formation of new cell adhesions, IGF-1R transiently relocalized to the plasma membrane from the Golgi. Thus, phosphorylation at Tyr1250/1251 promoted IGF-1R translocation to and signaling from the Golgi to support an aggressive cancer phenotype. This process distinguishes IGF-1R from IR signaling and could contribute to the poor clinical efficacy of antibodies that target IGF-1R on the cell surface.

A Golgi-targeted off-on fluorescent probe for real-time monitoring of pH changes in vivo.

We present the first Golgi-targeted small-molecular pH-sensitive fluorescent probe RSG, which allows an off-on fluorescence response to Golgi acidification with high sensitivity and specificity. RSG has been successfully used for real-time monitoring of Golgi pH changes induced by drug treatment at the cellular level, as well as by the LPS-mediated inflammation in vivo.

Golgi Apparatus Polarity Indicates Depression-Like Behaviors of Mice Using in Vivo Fluorescence Imaging.

Depression is associated with decreased expression of brain-derived neurotrophic factor (BDNF) which assembled in Golgi apparatus. The changes might be closely related to variation in Golgi apparatus polarity. Thus, developing a nondestructive method to detect polarity in living cells and in vivo can facilitate accurate diagnosis and prognosis of depression. Herein, we created a new near-infrared Golgi-targetable fluorescent probe (Golgi-P) in which the merocyanine and benzoyl difluoroboronate moieties sense polarity changes. Golgi-P exhibited a decrease in fluorescence intensity and red-shift of maximum emission wavelength as the increase in polarity. Using Golgi-P, we discovered distinctly higher polarity in brains of mice with depression phenotype for the first time. Furthermore, our results disclosed that the elevation of polarity could due to the reduced synthesis of BDNF. Altogether, this study offers a new strategy for the accurate diagnosis of depression.

The von Willebrand factor D'D3 assembly and structural principles for factor VIII binding and concatemer biogenesis.

D assemblies make up half of the von Willebrand factor (VWF), yet are of unknown structure. D1 and D2 in the prodomain and D'D3 in mature VWF at Golgi pH form helical VWF tubules in Weibel Palade bodies and template dimerization of D3 through disulfides to form ultralong VWF concatemers. D'D3 forms the binding site for factor VIII. The crystal structure of monomeric D'D3 with cysteine residues required for dimerization mutated to alanine was determined at an endoplasmic reticulum (ER)-like pH. The smaller C8-3, TIL3 (trypsin inhibitor-like 3), and E3 modules pack through specific interfaces as they wind around the larger, N-terminal, Ca2+-binding von Willebrand D domain (VWD) 3 module to form a wedge shape. D' with its TIL' and E' modules projects away from D3. The 2 mutated cysteines implicated in D3 dimerization are buried, providing a mechanism for protecting them against premature disulfide linkage in the ER, where intrachain disulfide linkages are formed. D3 dimerization requires co-association with D1 and D2, Ca2+, and Golgi-like acidic pH. Associated structural rearrangements in the C8-3 and TIL3 modules are required to expose cysteine residues for disulfide linkage. Our structure provides insight into many von Willebrand disease mutations, including those that diminish factor VIII binding, which suggest that factor VIII binds not only to the N-terminal TIL' domain of D' distal from D3 but also extends across 1 side of D3. The organizing principle for the D3 assembly has implications for other D assemblies and the construction of higher-order, disulfide-linked assemblies in the Golgi in both VWF and mucins.

Lysine at position 329 within a C-terminal dilysine motif is crucial for the ER localization of human SLC35B4.

SLC35B4 belongs to the solute carrier 35 (SLC35) family whose best-characterized members display a nucleotide sugar transporting activity. Using an experimental model of HepG2 cells and indirect immunofluorescent staining, we verified that SLC35B4 was localized to the endoplasmic reticulum (ER). We demonstrated that dilysine motif, especially lysine at position 329, is crucial for the ER localization of this protein in human cells and therefore one should use protein C-tagging with caution. To verify the importance of the protein in glycoconjugates synthesis, we generated SLC35B4-deficient HepG2 cell line using CRISPR-Cas9 approach. Our data showed that knock-out of the SLC35B4 gene does not affect major UDP-Xyl- and UDP-GlcNAc-dependent glycosylation pathways.

Identification and characterisation of a cryptic Golgi complex in Naegleria gruberi.

Although the Golgi complex has a conserved morphology of flattened stacked cisternae in most eukaryotes, it has lost the stacked organisation in several lineages, raising the question of what range of morphologies is possible for the Golgi. In order to understand this diversity, it is necessary to characterise the Golgi in many different lineages. Here, we identify the Golgi complex in Naegleria, one of the first descriptions of an unstacked Golgi organelle in a non-parasitic eukaryote, other than fungi. We provide a comprehensive list of Golgi-associated membrane trafficking genes encoded in two species of Naegleria and show that nearly all are expressed in mouse-passaged N. fowleri cells. We then study distribution of the Golgi marker (Ng)CopB by fluorescence in Naegleria gruberi, identifying membranous structures that are disrupted by Brefeldin A treatment, consistent with Golgi localisation. Confocal and immunoelectron microscopy reveals that NgCOPB localises to tubular membranous structures. Our data identify the Golgi organelle for the first time in this major eukaryotic lineage, and provide the rare example of a tubular morphology, representing an important sampling point for the comparative understanding of Golgi organellar diversity.This article has an associated First Person interview with the first author of the paper.

TANGO1 builds a machine for collagen export by recruiting and spatially organizing COPII, tethers and membranes.

Collagen export from the endoplasmic reticulum (ER) requires TANGO1, COPII coats, and retrograde fusion of ERGIC membranes. How do these components come together to produce a transport carrier commensurate with the bulky cargo collagen? TANGO1 is known to form a ring that corrals COPII coats, and we show here how this ring or fence is assembled. Our data reveal that a TANGO1 ring is organized by its radial interaction with COPII, and lateral interactions with cTAGE5, TANGO1-short or itself. Of particular interest is the finding that TANGO1 recruits ERGIC membranes for collagen export via the NRZ (NBAS/RINT1/ZW10) tether complex. Therefore, TANGO1 couples retrograde membrane flow to anterograde cargo transport. Without the NRZ complex, the TANGO1 ring does not assemble, suggesting its role in nucleating or stabilising this process. Thus, coordinated capture of COPII coats, cTAGE5, TANGO1-short, and tethers by TANGO1 assembles a collagen export machine at the ER.

isGPT: An optimized model to identify sub-Golgi protein types using SVM and Random Forest based feature selection.

The Golgi Apparatus (GA) is a key organelle for protein synthesis within the eukaryotic cell. The main task of GA is to modify and sort proteins for transport throughout the cell. Proteins permeate through the GA on the ER (Endoplasmic Reticulum) facing side (cis side) and depart on the other side (trans side). Based on this phenomenon, we get two types of GA proteins, namely, cis-Golgi protein and trans-Golgi protein. Any dysfunction of GA proteins can result in congenital glycosylation disorders and some other forms of difficulties that may lead to neurodegenerative and inherited diseases like diabetes, cancer and cystic fibrosis. So, the exact classification of GA proteins may contribute to drug development which will further help in medication. In this paper, we focus on building a new computational model that not only introduces easy ways to extract features from protein sequences but also optimizes classification of trans-Golgi and cis-Golgi proteins. After feature extraction, we have employed Random Forest (RF) model to rank the features based on the importance score obtained from it. After selecting the top ranked features, we have applied Support Vector Machine (SVM) to classify the sub-Golgi proteins. We have trained regression model as well as classification model and found the former to be superior. The model shows improved performance over all previous methods. As the benchmark dataset is significantly imbalanced, we have applied Synthetic Minority Over-sampling Technique (SMOTE) to the dataset to make it balanced and have conducted experiments on both versions. Our method, namely, identification of sub-Golgi Protein Types (isGPT), achieves accuracy values of 95.4%, 95.9% and 95.3% for 10-fold cross-validation test, jackknife test and independent test respectively. According to different performance metrics, isGPT performs better than state-of-the-art techniques. The source code of isGPT, along with relevant dataset and detailed experimental results, can be found at https://github.com/srautonu/isGPT.

Proteomics Identifies Golgi phosphoprotein 3 (GOLPH3) with A Link Between Golgi Structure, Cancer, DNA Damage and Protection from Cell Death.

GOLPH3 is the first example of a Golgi resident oncogene protein. It was independently identified in multiple screens; first in proteomic-based screens as a resident protein of the Golgi apparatus, and second as an oncogene product in a screen for genes amplified in cancer. A third screen uncovered the association of GOLPH3 with the Golgi resident phospholipid, phosphatidyl inositol 4 phosphate (PI4P) to maintain the characteristic ribbon structure of the Golgi apparatus favoring vesicular transport of secretory proteins.

A smart drug: a pH-responsive photothermal ablation agent for Golgi apparatus activated cancer therapy.

We report a pH-responsive photothermal ablation agent (pH-PTT) based on cyanine dyes for photothermal therapy (PTT). The nanoparticles formed by BSA and pH-PTT preferentially accumulated in the Golgi apparatus of cancer cells compared to normal cells, and thus can be specifically activated by the acidic Golgi apparatus in cancer cells for effective PTT both ex vivo and in vivo.

Mycobacterium tuberculosis Infection Manipulates the Glycosylation Machinery and the N-Glycoproteome of Human Macrophages and Their Microparticles.

Tuberculosis (TB) remains a prevalent and lethal infectious disease. The glycobiology associated with Mycobacterium tuberculosis infection of frontline alveolar macrophages is still unresolved. Herein, we investigated the regulation of protein N-glycosylation in human macrophages and their secreted microparticles (MPs) used for intercellular communication upon M. tb infection. LC-MS/MS-based proteomics and glycomics were performed to monitor the regulation of glycosylation enzymes and receptors and the N-glycome in in vitro-differentiated macrophages and in isolated MPs upon M. tb infection. Infection promoted a dramatic regulation of the macrophage proteome. Most notably, significant infection-dependent down-regulation (4-26 fold) of 11 lysosomal exoglycosidases, e.g., ß-galactosidase, ß-hexosaminidases and α-/ß-mannosidases, was observed. Relative weak infection-driven transcriptional regulation of these exoglycosidases and a stronger augmentation of the extracellular hexosaminidase activity demonstrated that the lysosome-centric changes may originate predominantly from infection-induced secretion of the lysosomal content. The macrophages showed heterogeneous N-glycan profiles and displayed significant up-regulation of complex-type glycosylation and concomitant down-regulation of paucimannosylation upon infection. Complementary intact N-glycopeptide analysis supported a subcellular-specific manipulation of the glycosylation machinery and altered glycosylation patterns of lysosomal N-glycoproteins within infected macrophages. Interestingly, the corresponding macrophage-derived MPs displayed unique N-glycome and proteome signatures supporting a preferential packaging from plasma membranes. The MPs were devoid of infection-dependent N-glycosylation signatures, but interestingly displayed increased levels of the glyco-initiating oligosaccharyltransferase complex and associated α-glucosidases that correlated with increased formation, N-glycan precursor levels and N-glycan density of infected MPs. In conclusion, this system-wide study provides new insight into the host- and pathogen-driven N-glycoproteome manipulation of macrophages in TB.

Nanoscale Distribution of Transforming Growth Factor Receptor on Post-Golgi Vesicle Revealed by Super-resolution Microscopy.

Transforming growth factor-ß (TGF-ß) type II receptor (TßRII) plays a critical role in the initiation of TGF-ß signaling pathway; therefore, the study of its synthesis and transport processes is of great important. In this work, we achieved super-resolution imaging of a new type of TßRII-containing post-Golgi vesicle by our home-built stimulated emission depletion (STED) microscope. We visualized the ring-shaped structure of these vesicles containing newly synthesized TßRII in the cytoplasm and characterized their size distribution from 300 to 1000 nm. These vesicles could be swollen by chloroquine treatment. Further investigation revealed that TßRII formed clusters on the outer ring of the post-Golgi vesicles. This study offers new information on the intracellular transportation of TGF-ß receptors for better understanding its signaling process.