Intracellular Localization of Processing Events in Human Surfactant Protein B Biosynthesis

Korimilli, Annapurna; Gonzales, Linda W.; Guttentag, Susan H.

doi:10.1074/jbc.275.12.8672

Cited by 65 publications

(63 citation statements)

References 45 publications

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“…But, this does not necessarily exclude the possibility that the Rab38 participates in specific steps in the transport of surfactant components. Because SP-A and SP-B are highly enriched in LB and also found throughout the secretory pathway, 22 the co-localization of these proteins with Rab38 is unlikely to be extensive.…”

Section: Discussionmentioning

confidence: 99%

“…No single cell expresses all of the rab family members. However, there are many Rab proteins whose expressions are ubiquitous over a variety of tissues and cells, such as Rab 1,2,4,5,[6][7][8][9][10][11][12][13][14]18,20,22,24,28, and 30. 2 Expression of some Rab proteins are highly regulated depending on cell type (epithelial, mesenchymal, or endothelial) and specific phenotype (differentiation, activation, or polarization).…”

Section: Discussionmentioning

confidence: 99%

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Expression and Localization of a Novel Rab Small G Protein (Rab38) in the Rat Lung

Osanai

Iguchi

Takahashi

et al. 2001

The American Journal of Pathology

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The Rab small G protein family participates in intracellular vesicle transport, including exocytosis and endocytosis. The cDNA encoding a novel Rab-related small G protein (Rab38) has been cloned from rat lung cDNA library and recorded in GenBank (accession no. M94043). However, the expression and localization of the protein in the lung remains primarily unknown. We produced polyhistidine-tagged recombinant Rab38 and a polyclonal antibody with a synthetic peptide. Immunohistochemistry demonstrated that the protein is specifically localized in alveolar type II cells and in bronchial epithelial cells. In situ hybridization using a digoxygenin-labeled RNA riboprobe clearly showed that the mRNA of the protein is localized in alveolar type II cells and bronchial epithelial cells, especially terminal airway epithelial cells. Western blot and reverse transcriptase-polymerase chain reaction showed distinct expression of the protein and mRNA in isolated alveolar type II cells, but not in alveolar macrophages. The native protein was predominantly hydrophobic and was enriched in a high-density vesicle fraction but was barely detectable in nuclear and lamellar body fractions in alveolar type II cells. Immunofluorescence cytochemistry performed on cultured alveolar type II cells showed that Rab38 distributed extensively in the cytoplasm with a distribution pattern similar to endoplasmic reticulum rather than other subcellular organelles. These results suggest that this novel rab small G protein (Rab38) mediates vesicular transport in terminal airway epithelium. Small GTP-binding proteins are a group of monomeric intracellular proteins that have GTP/GDP binding and GTPase activities, and mediate essential cell functions, including cell growth/differentiation, cytoskeletal configuration (cell movement, change of shape, and contraction and relaxation), and intracellular vesicle transport, including exocytosis and endocytosis. 1,2 The ras p21 was found first and was soon followed by several other proteins. Now, more than 50 members form a ras superfamily. They have highly conserved domains, contributing to interaction with guanine nucleotides, in organisms from yeast through mammals. Most of these proteins are prenylated at their carboxy termini and specific subsets are also modified by palmitoylation. The ras-related superfamily is now recognized as consisting of three major gene families (ras, rho, and rab) and other minor families.The rab family has more than 30 members, the largest number among small G proteins. The Rab proteins are known to localize to specific cell organelles, and are found in both membrane-bound and cytosolic forms. 2,3 Thus, they are believed to mediate intracellular vesicle transport among restricted intracellular compartments. Although current information in the sequence database indicates more than 30 members, there are few Rab proteins for which intracellular localization and function have been clarified.A novel cDNA has been cloned from the rat lung cDNA library encoding a rab-related small G protei...

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Expression and Localization of a Novel Rab Small G Protein (Rab38) in the Rat Lung

Osanai

Iguchi

Takahashi

et al. 2001

The American Journal of Pathology

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“…The polyclonal antibody against napsin A was produced by immunizing rabbits with a peptide of 15 amino acids (SFYLNRDPEEPDGGE) as previously described (14). The C-terminal proSP-B peptide sequence Gly 284 -Ser 304 (CFlankSP-B) was chosen for production of synthetic peptides and antiserum preparation as described previously (7,8). Rabbit antiserum and a monoclonal antibody specific for mature SP-B were kindly donated by Dr. J.…”

Section: Methodsmentioning

confidence: 99%

“…On route from its site of synthesis to the lamellar bodies, the processing to mature SP-B involves the cleavage of the signal peptide, glycosylation of the C terminus, followed by the cleavage of the N-terminal and C-terminal propeptide (7). The processing of the ϳ42-kDa proSP-B through ϳ23-25-and ϳ9-kDa proSP-B processing intermediates to mature 8-kDa SP-B is an at least three-step process with two distinct cleavages of the N-terminal propeptide and one of the C-terminal propeptide (8). Although various proSP-B processing steps and the site of proSP-B processing have been described (8,9), little is known about the identity of the proteases involved in the posttranslational processing.…”

mentioning

confidence: 99%

“…The processing of the ϳ42-kDa proSP-B through ϳ23-25-and ϳ9-kDa proSP-B processing intermediates to mature 8-kDa SP-B is an at least three-step process with two distinct cleavages of the N-terminal propeptide and one of the C-terminal propeptide (8). Although various proSP-B processing steps and the site of proSP-B processing have been described (8,9), little is known about the identity of the proteases involved in the posttranslational processing.…”

mentioning

confidence: 99%

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Involvement of Napsin A in the C- and N-terminal Processing of Surfactant Protein B in Type-II Pneumocytes of the Human Lung

Brasch

Ochs

Kähne

et al. 2003

Journal of Biological Chemistry

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Surfactant protein B (SP-B) is a critical component of pulmonary surfactant, and a deficiency of active SP-B results in fatal respiratory failure. SP-B is synthesized by type-II pneumocytes as a 42-kDa propeptide (proSP-B), which is posttranslationally processed to an 8-kDa surface-active protein. Napsin A is an aspartic protease expressed in type-II pneumocytes. To characterize the role of napsin A in the processing of proSP-B, we colocalized napsin A and precursors of SP-B as well as SP-B in the Golgi complex, multivesicular, composite, and lamellar bodies of type-II pneumocytes in human lungs using immunogold labeling. Furthermore, we measured aspartic protease activity in isolated lamellar bodies as well as isolated human type-II pneumocytes and studied the cleavage of proSP-B by napsin A and isolated lamellar bodies in vitro. Both, napsin A and isolated lamellar bodies cleaved proSP-B and generated three identical processing products. Processing of proSP-B by isolated lamellar bodies was completely inhibited by an aspartic protease inhibitor. Sequence analysis of proSP-B processing products revealed several cleavage sites in the Nand C-terminal propeptides as well as one in the mature peptide. Two of the four processing products generated in vitro were also detected in type-II pneumocytes. In conclusion, our results show that napsin A is involved in the N-and C-terminal processing of proSP-B in type-II pneumocytes.The integrity and function of pulmonary surfactant are of paramount importance for lung function. Disturbance of surfactant activity leads to respiratory distress (1). The main function of pulmonary surfactant is the reduction of the surface tension at the air/liquid interface in the lung, thus preventing alveolar collapse at end-expiration. Pulmonary surfactant is a complex mixture of ϳ90% lipids and ϳ10% proteins that is synthesized, stored, secreted, and to a large extent recycled by type-II pneumocytes of the alveolar epithelium.The hydrophobic surfactant protein B (SP-B) 1 interacts with phospholipids and contributes to the formation of intracellular lamellar bodies, the structural rearrangement of secreted surfactant lipids into tubular myelin, as well as the subsequent rapid insertion of secreted surfactant phospholipids into the surface film (reviewed in Ref. 2). Hereditary SP-B deficiency in infants or mice leads to respiratory failure at birth (3-6). However, hereditary alveolar proteinosis in babies without any detectable mutations in the SP-B gene as well as acquired pulmonary alveolar proteinosis in children and adults are characterized by an intraalveolar accumulation of mature surfactant proteins and abnormal SP-B precursors. Furthermore, only SP-B precursors are detected in babies with congenital surfactant defects characterized by the absence of lamellar bodies in type-II pneumocytes.2 Therefore, insufficient processing of proSP-B due to a lack or dysfunction of one or more proteases involved in SP-B processing might be yet another undiscovered cause of surfactant dysfunction in p...

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Microstructured Hydrogels to Guide Self‐Assembly and Function of Lung Alveolospheres

et al. 2022

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Epithelial cell organoids have increased opportunities to probe questions on tissue development and disease in vitro and for therapeutic cell transplantation. Despite their potential, current protocols to grow these organoids almost exclusively depend on culture within 3D Matrigel, which limits defined culture conditions, introduces animal components, and results in heterogenous organoids (i.e., shape, size, composition). Here, a method is described that relies on hyaluronic acid hydrogels for the generation and expansion of lung alveolar organoids (alveolospheres). Using synthetic hydrogels with defined chemical and physical properties, human‐induced pluripotent stem cell (iPSC)‐derived alveolar type 2 cells (iAT2s) self‐assemble into alveolospheres and propagate in Matrigel‐free conditions. By engineering predefined microcavities within these hydrogels, the heterogeneity of alveolosphere size and structure is reduced when compared to 3D culture, while maintaining the alveolar type 2 cell fate of human iAT2‐derived progenitor cells. This hydrogel system is a facile and accessible system for the culture of iPSC‐derived lung progenitors and the method can be expanded to the culture of primary mouse tissue derived AT2 and other epithelial progenitor and stem cell aggregates.

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Intracellular Localization of Processing Events in Human Surfactant Protein B Biosynthesis

Cited by 65 publications

References 45 publications

Expression and Localization of a Novel Rab Small G Protein (Rab38) in the Rat Lung

Expression and Localization of a Novel Rab Small G Protein (Rab38) in the Rat Lung

Involvement of Napsin A in the C- and N-terminal Processing of Surfactant Protein B in Type-II Pneumocytes of the Human Lung

Microstructured Hydrogels to Guide Self‐Assembly and Function of Lung Alveolospheres

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