ABSTRACT
In an adenosine triphosphate (ATP)-dependent process, the hSWI/SNF chromatin remodeling complex functions to alter chromatin structure, thereby regulating transcription factor access to DNA. In addition to interactions with transcription factors and recognition of acetylated histone residues, the chromatin remodeling activity of hSWI/SNF has also been shown to respond to a variety of cell signaling pathways. Our results demonstrate a novel interaction between the serine/threonine kinase Akt and members of the hSWI/SNF chromatin remodeling complex. Activation of Akt in HeLa cells resulted in its association with hSWI/SNF subunits: INI1, BAF155 and BAF170, as well as actin. BAF155 became preferentially recognized by an antibody that detects phosphorylated Akt substrates upon activation of Akt, suggesting that BAF155 may be an in vivo target for phosphorylation by Akt. Glutathione-S-transferase (GST) pulldown experiments demonstrated that INI1 and BAF155 were both capable of directly interacting with Akt. Finally, in vitro kinase assays provided additional evidence that BAF155 and potentially INI1 are substrates for Akt phosphorylation. These data provide the first evidence that Akt signaling may modulate function of the hSWI/SNF complex.
Subject(s)
Chromatin/chemistry , Proto-Oncogene Proteins c-akt/metabolism , Actins/metabolism , Adenosine Triphosphate/metabolism , Basic Helix-Loop-Helix Leucine Zipper Transcription Factors/metabolism , Chromatin/metabolism , Chromosomal Proteins, Non-Histone , Cytoplasm/metabolism , DNA-Binding Proteins/metabolism , Erythroid-Specific DNA-Binding Factors/metabolism , HeLa Cells , Humans , Phosphorylation , Protein Binding , SMARCB1 Protein , Transcription Factors/metabolismABSTRACT
The Harderian gland (HG), a sero-mucous secreting organ in the eye orbit, has long been recognized as immunologically important in chickens. During experimentation to characterize immune components of the gland, proteomics analysis revealed the presence of hematopoietic prostaglandin D synthase (H-PGDS). Extraction of total RNA followed by RT-PCR produced cDNA of 597 base pairs. DNA sequencing revealed nucleic acid and predicted amino acid sequences that were 99% aligned with the one published sequence for chicken H-PGDS of the spleen. Alignment with murine, rat, and human H-PGDS were 69, 69, and 66%, respectively. Ocular vaccination of chickens with a Newcastle Disease/Infectious Bronchitis vaccine (Mass.-Ark. Strain) induced an increase in H-PGDS expression determined by real-time PCR. Furthermore, immunohistochemistry of frozen HG sections showed positive stained cells for both H-PGDS and mast cell tryptase in the sub-epithelial cell layers of the HG ducts. Based on the potent vasoactive role of PGD(2), it appears that the chicken HG is a site of active mucosal immunity partially mediated by PGD(2) synthesized by H-PGDS in the gland.
