Role of FQQI motif in the internalization, trafficking, and signaling of guanylyl-cyclase/natriuretic peptide receptor-A in cultured murine mesangial cells.
Am J Physiol Renal Physiol
; 310(1): F68-84, 2016 Jan 01.
Article
in En
| MEDLINE
| ID: mdl-26377794
ABSTRACT
Binding of the cardiac hormone atrial natriuretic peptide (ANP) to transmembrane guanylyl cyclase/natriuretic peptide receptor-A (GC-A/NPRA), produces the intracellular second messenger cGMP in target cells. To delineate the critical role of an endocytic signal in intracellular sorting of the receptor, we have identified a FQQI (Phe(790), Gln(791), Gln(792), and Ile(793)) motif in the carboxyl-terminal region of NPRA. Mouse mesangial cells (MMCs) were transiently transfected with the enhanced green fluorescence protein (eGFP)-tagged wild-type (WT) and mutant constructs of eGFP-NPRA. The mutation FQQI/AAAA, in the eGFP-NPRA cDNA sequence, markedly attenuated the internalization of mutant receptors by almost 49% compared with the WT receptor. Interestingly, we show that the µ1B subunit of adaptor protein-1 binds directly to a phenylalanine-based FQQI motif in the cytoplasmic tail of the receptor. However, subcellular trafficking indicated that immunofluorescence colocalization of the mutated receptor with early endosome antigen-1 (EEA-1), lysosome-associated membrane protein-1 (LAMP-1), and Rab 11 marker was decreased by 57% in early endosomes, 48% in lysosomes, and 42% in recycling endosomes, respectively, compared with the WT receptor in MMCs. The receptor containing the mutated motif (FQQI/AAAA) also produced a significantly decreased level of intracellular cGMP during subcellular trafficking than the WT receptor. The coimmunoprecipitation assay confirmed a decreased level of colocalization of the mutant receptor with subcellular compartments during endocytic processes. The results suggest that the FQQI motif is essential for the internalization and subcellular trafficking of NPRA during the hormone signaling process in intact MMCs.
Key words
Full text:
1
Collection:
01-internacional
Database:
MEDLINE
Main subject:
Second Messenger Systems
/
Receptors, Atrial Natriuretic Factor
/
Endocytosis
/
Mesangial Cells
Type of study:
Prognostic_studies
Limits:
Animals
Language:
En
Journal:
Am J Physiol Renal Physiol
Journal subject:
FISIOLOGIA
/
NEFROLOGIA
Year:
2016
Document type:
Article