Signaling from the Secretory Granule to the Nucleus: Uhmk1 and PAM

• Published in: Molecular endocrinology (Baltimore, Md.) Vol. 24; no. 8; pp. 1543 - 1558
• Main Authors: Francone, Victor P, Ifrim, Marius F, Rajagopal, Chitra, Leddy, Christopher J, Wang, Yanping, Carson, John H, Mains, Richard E, Eipper, Betty A
• Format: Journal Article
• Language: English
• Published: United States Endocrine Society 01.08.2010; Oxford University Press; The Endocrine Society
• Subjects: Animals; Aquaporin 1 - genetics; Aquaporin 1 - metabolism; Blotting, Western; Cell Line, Tumor; Cell Nucleus - metabolism; Cells, Cultured; Cytoplasm - metabolism; Flow Cytometry; Fluorescence Recovery After Photobleaching; Immunohistochemistry; Intracellular Signaling Peptides and Proteins - genetics; Intracellular Signaling Peptides and Proteins - metabolism; Mice; Microscopy, Fluorescence; Mixed Function Oxygenases - genetics; Mixed Function Oxygenases - metabolism; Multienzyme Complexes - genetics; Multienzyme Complexes - metabolism; Oligonucleotide Array Sequence Analysis; Polymerase Chain Reaction; Protein-Serine-Threonine Kinases - genetics; Protein-Serine-Threonine Kinases - metabolism; Rats; Secretory Leukocyte Peptidase Inhibitor - genetics; Secretory Leukocyte Peptidase Inhibitor - metabolism; Secretory Vesicles - metabolism
• ISSN: 0888-8809; 1944-9917
• DOI: 10.1210/me.2009-0381

Neurons and endocrine cells package peptides in secretory granules (large dense-core vesicles) for storage and stimulated release. Studies of peptidylglycine α-amidating monooxygenase (PAM), an essential secretory granule membrane enzyme, revealed a pathway that can relay information from secretory granules to the nucleus, resulting in alterations in gene expression. The cytosolic domain (CD) of PAM, a type 1 membrane enzyme essential for the production of amidated peptides, is basally phosphorylated by U2AF homology motif kinase 1 (Uhmk1) and other Ser/Thr kinases. Proopiomelanocortin processing in AtT-20 corticotrope tumor cells was increased when Uhmk1 expression was reduced. Uhmk1 was concentrated in the nucleus, but cycled rapidly between nucleus and cytosol. Endoproteolytic cleavage of PAM releases a soluble CD fragment that localizes to the nucleus. Localization of PAM-CD to the nucleus was decreased when PAM-CD with phosphomimetic mutations was examined and when active Uhmk1 was simultaneously overexpressed. Membrane-tethering Uhmk1 did not eliminate its ability to exclude PAM-CD from the nucleus, suggesting that cytosolic Uhmk1 could cause this response. Microarray analysis demonstrated the ability of PAM to increase expression of a small subset of genes, including aquaporin 1 (Aqp1) in AtT-20 cells. Aqp1 mRNA levels were higher in wild-type mice than in mice heterozygous for PAM, indicating that a similar relationship occurs in vivo. Expression of PAM-CD also increased Aqp1 levels whereas expression of Uhmk1 diminished Aqp1 expression. The outlines of a pathway that ties secretory granule metabolism to the transcriptome are thus apparent. A pathway through which Uhmk1 phosphorylation of the cytosolic domain of the peptide amidating enzyme links secretory granule metabolism to the transcriptome is identified.