Limits...
Porosome in Cystic Fibrosis.

Jena BP - Discoveries (Craiova) (2014 Jul-Sep)

Bottom Line: This understanding now provides a platform to address diseases that may result from secretory defects.Hence secretion of more viscous mucus prevents its proper transport, resulting in chronic and fatal airways disease such as cystic fibrosis (CF).The involvement of CFTR in porosome-mediated mucin secretion is hypothesized, and is currently being tested.

View Article: PubMed Central - PubMed

Affiliation: Wayne State University School of Medicine, Department of Physiology, Detroit, MI, USA.

ABSTRACT

Macromolecular structures embedded in the cell plasma membrane called 'porosomes', are involved in the regulated fractional release of intravesicular contents from cells during secretion. Porosomes range in size from 15 nm in neurons and astrocytes to 100-180 nm in the exocrine pancreas and neuroendocrine cells. Porosomes have been isolated from a number of cells, and their morphology, composition, and functional reconstitution well documented. The 3D contour map of the assembly of proteins within the porosome complex, and its native X-ray solution structure at sub-nm resolution has also advanced. This understanding now provides a platform to address diseases that may result from secretory defects. Water and ion binding to mucin impart hydration, critical for regulating viscosity of the mucus in the airways epithelia. Appropriate viscosity is required for the movement of mucus by the underlying cilia. Hence secretion of more viscous mucus prevents its proper transport, resulting in chronic and fatal airways disease such as cystic fibrosis (CF). CF is caused by the malfunction of CF transmembrane conductance regulator (CFTR), a chloride channel transporter, resulting in viscous mucus in the airways. Studies in mice lacking functional CFTR secrete highly viscous mucous that adhered to the epithelium. Since CFTR is known to interact with the t-SNARE protein syntaxin-1A, and with the chloride channel CLC-3, which are also components of the porosome complex, the interactions between CFTR and the porosome complex in the mucin-secreting human airway epithelial cell line Calu-3 was hypothesized and tested. Results from the study demonstrate the presence of approximately 100 nm in size porosome complex composed of 34 proteins at the cell plasma membrane in Calu-3 cells, and the association of CFTR with the complex. In comparison, the nuclear pore complex measures 120 nm and is comprised of over 500 protein molecules. The involvement of CFTR in porosome-mediated mucin secretion is hypothesized, and is currently being tested.

No MeSH data available.


Related in: MedlinePlus

Schematic drawing depicting the presence and increased association of dynamin with the porosome complex following stimulation of neurotransmitter release12, which may be similar in the mucin-secreting Calu-3 cell. Following stimulation of secretion, synaptic vesicles would dock at the porosome base, develop intravesicular pressure via active transport of water through water channels or aquaporins (AQP) at the vesicle membrane, transiently fuse at the porosome base via SNAREs and calcium, and expel neurotransmitters. After secretion, NSF an ATPase, and dynamin a GTPase, would work synchronously to disassembly t-/v-SNARE complexes and fission the neck of fused vesicles at the porosome base respectively. By this mechanism, partially empty vesicles could go through multiple rounds of docking-fusion-expulsion-dissociation. Unlike protein and peptide containing vesicles, synaptic vesicles have neurotransmitter transporters at the vesicle membrane to rapidly refill vesicles12. In case of the Calu-3 cell, once mucin containing vesicles empty, they may recycle via the endosome or lysosomal pathway. ©Bhanu Jena.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4581455&req=5

Figure 6: Schematic drawing depicting the presence and increased association of dynamin with the porosome complex following stimulation of neurotransmitter release12, which may be similar in the mucin-secreting Calu-3 cell. Following stimulation of secretion, synaptic vesicles would dock at the porosome base, develop intravesicular pressure via active transport of water through water channels or aquaporins (AQP) at the vesicle membrane, transiently fuse at the porosome base via SNAREs and calcium, and expel neurotransmitters. After secretion, NSF an ATPase, and dynamin a GTPase, would work synchronously to disassembly t-/v-SNARE complexes and fission the neck of fused vesicles at the porosome base respectively. By this mechanism, partially empty vesicles could go through multiple rounds of docking-fusion-expulsion-dissociation. Unlike protein and peptide containing vesicles, synaptic vesicles have neurotransmitter transporters at the vesicle membrane to rapidly refill vesicles12. In case of the Calu-3 cell, once mucin containing vesicles empty, they may recycle via the endosome or lysosomal pathway. ©Bhanu Jena.

Mentions: New and recently developed crosslinkers79 combined with tandem mass spectrometry are being carried out, which will provide identities of interacting subunits and provide the identities of specific residues crosslinked both between and within subunits in the porosome complex. Results from these studies will provide information on interaction domains and distance constraints on protein structures. Quantitative mass spectrometry using iTRAQ are also being carried out, which will provide additional information on changes in porosome subunits composition and dynamics, as a function of the secretion status of the organelle. Immuno-AFM5, immuno–EM, and SAXS80 (Figure 5) studies on isolated Calu-3 porosomes as in porosomes of the exocrine pancreas and neurons, are being performed to determine the distribution of some of the major proteins within the complex. Similar to studies using SRM on the nuclear pore complex81, SRM is being employed to obtain additional information on the structure of the mucin-secreting porosome complex. Finally, computational approaches are being employed, such as coarse-grain molecular docking studies82–97, homology modeled interactions98–100, and fitting of known atomic structures of protein-protein interactions and complexes101–108. It is becoming increasingly clear that the ultrastructural and mass spectrometry methods show promise in providing complementary information and the high degree of cross-validation required to build an accurate structural model of the mucin-secreting porosome complex. Collectively, the outlined studies briefly discussed here will enable an understanding at the molecular level, the elegant mechanism of porosome-mediated secretion (Figure 6) in Calu-3 and other cells.


Porosome in Cystic Fibrosis.

Jena BP - Discoveries (Craiova) (2014 Jul-Sep)

Schematic drawing depicting the presence and increased association of dynamin with the porosome complex following stimulation of neurotransmitter release12, which may be similar in the mucin-secreting Calu-3 cell. Following stimulation of secretion, synaptic vesicles would dock at the porosome base, develop intravesicular pressure via active transport of water through water channels or aquaporins (AQP) at the vesicle membrane, transiently fuse at the porosome base via SNAREs and calcium, and expel neurotransmitters. After secretion, NSF an ATPase, and dynamin a GTPase, would work synchronously to disassembly t-/v-SNARE complexes and fission the neck of fused vesicles at the porosome base respectively. By this mechanism, partially empty vesicles could go through multiple rounds of docking-fusion-expulsion-dissociation. Unlike protein and peptide containing vesicles, synaptic vesicles have neurotransmitter transporters at the vesicle membrane to rapidly refill vesicles12. In case of the Calu-3 cell, once mucin containing vesicles empty, they may recycle via the endosome or lysosomal pathway. ©Bhanu Jena.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4581455&req=5

Figure 6: Schematic drawing depicting the presence and increased association of dynamin with the porosome complex following stimulation of neurotransmitter release12, which may be similar in the mucin-secreting Calu-3 cell. Following stimulation of secretion, synaptic vesicles would dock at the porosome base, develop intravesicular pressure via active transport of water through water channels or aquaporins (AQP) at the vesicle membrane, transiently fuse at the porosome base via SNAREs and calcium, and expel neurotransmitters. After secretion, NSF an ATPase, and dynamin a GTPase, would work synchronously to disassembly t-/v-SNARE complexes and fission the neck of fused vesicles at the porosome base respectively. By this mechanism, partially empty vesicles could go through multiple rounds of docking-fusion-expulsion-dissociation. Unlike protein and peptide containing vesicles, synaptic vesicles have neurotransmitter transporters at the vesicle membrane to rapidly refill vesicles12. In case of the Calu-3 cell, once mucin containing vesicles empty, they may recycle via the endosome or lysosomal pathway. ©Bhanu Jena.
Mentions: New and recently developed crosslinkers79 combined with tandem mass spectrometry are being carried out, which will provide identities of interacting subunits and provide the identities of specific residues crosslinked both between and within subunits in the porosome complex. Results from these studies will provide information on interaction domains and distance constraints on protein structures. Quantitative mass spectrometry using iTRAQ are also being carried out, which will provide additional information on changes in porosome subunits composition and dynamics, as a function of the secretion status of the organelle. Immuno-AFM5, immuno–EM, and SAXS80 (Figure 5) studies on isolated Calu-3 porosomes as in porosomes of the exocrine pancreas and neurons, are being performed to determine the distribution of some of the major proteins within the complex. Similar to studies using SRM on the nuclear pore complex81, SRM is being employed to obtain additional information on the structure of the mucin-secreting porosome complex. Finally, computational approaches are being employed, such as coarse-grain molecular docking studies82–97, homology modeled interactions98–100, and fitting of known atomic structures of protein-protein interactions and complexes101–108. It is becoming increasingly clear that the ultrastructural and mass spectrometry methods show promise in providing complementary information and the high degree of cross-validation required to build an accurate structural model of the mucin-secreting porosome complex. Collectively, the outlined studies briefly discussed here will enable an understanding at the molecular level, the elegant mechanism of porosome-mediated secretion (Figure 6) in Calu-3 and other cells.

Bottom Line: This understanding now provides a platform to address diseases that may result from secretory defects.Hence secretion of more viscous mucus prevents its proper transport, resulting in chronic and fatal airways disease such as cystic fibrosis (CF).The involvement of CFTR in porosome-mediated mucin secretion is hypothesized, and is currently being tested.

View Article: PubMed Central - PubMed

Affiliation: Wayne State University School of Medicine, Department of Physiology, Detroit, MI, USA.

ABSTRACT

Macromolecular structures embedded in the cell plasma membrane called 'porosomes', are involved in the regulated fractional release of intravesicular contents from cells during secretion. Porosomes range in size from 15 nm in neurons and astrocytes to 100-180 nm in the exocrine pancreas and neuroendocrine cells. Porosomes have been isolated from a number of cells, and their morphology, composition, and functional reconstitution well documented. The 3D contour map of the assembly of proteins within the porosome complex, and its native X-ray solution structure at sub-nm resolution has also advanced. This understanding now provides a platform to address diseases that may result from secretory defects. Water and ion binding to mucin impart hydration, critical for regulating viscosity of the mucus in the airways epithelia. Appropriate viscosity is required for the movement of mucus by the underlying cilia. Hence secretion of more viscous mucus prevents its proper transport, resulting in chronic and fatal airways disease such as cystic fibrosis (CF). CF is caused by the malfunction of CF transmembrane conductance regulator (CFTR), a chloride channel transporter, resulting in viscous mucus in the airways. Studies in mice lacking functional CFTR secrete highly viscous mucous that adhered to the epithelium. Since CFTR is known to interact with the t-SNARE protein syntaxin-1A, and with the chloride channel CLC-3, which are also components of the porosome complex, the interactions between CFTR and the porosome complex in the mucin-secreting human airway epithelial cell line Calu-3 was hypothesized and tested. Results from the study demonstrate the presence of approximately 100 nm in size porosome complex composed of 34 proteins at the cell plasma membrane in Calu-3 cells, and the association of CFTR with the complex. In comparison, the nuclear pore complex measures 120 nm and is comprised of over 500 protein molecules. The involvement of CFTR in porosome-mediated mucin secretion is hypothesized, and is currently being tested.

No MeSH data available.


Related in: MedlinePlus