Immunohistochemical staining patterns using epitope-specific antibodies indicate conformation variants of antisecretory factor/S5a in the CNS.
Antisecretory factor (AF/S5a/Rpn10) was originally identified through its ability to counteract pathological secretion. AF is also a potent anti-inflammatory agent, a neuromodulator, and an important component of the proteasome. Human AF has a calculated molecular mass of 41 kDa and a pI of 4.7. No family of AF-like proteins has been identified. AF has multiple functions in the cell, and different functional forms could exist as a result of post-translational modifications. Epitope-specific antibodies covering the entire length of AF were used to investigate whether modified forms of AF could be detected in the porcine spinal cord by Western blots, 2D gels, and immunohistochemistry (IHC). Western blot and 2D gels showed that all antisera detected a single protein with very similar molecular mass and pI. However, IHC resulted in an epitope-specific subcellular staining pattern. Antisera recognizing epitopes in the N-terminal part of AF, containing the antisecretory activity, showed a more restricted localisation than antisera directed at the C-terminal part, containing the ubiquitin-binding sites. We suggest that AF can exist in several conformational variants, perhaps due to differences in redox state and/or pH in the various cellular compartments. Such conformational changes could be of functional importance.
Jennische
Eva
E
Institute of Anatomy and Cell Biology, Göteborg University, Göteborg, Sweden. eva.jennische@anatcell.gu.se
Johansson
Ewa
E
Hansson
Hans-Arne
HA
Jonson
Ingela
I
D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies / Q000134:cerebrospinal fluid; D002490:Central Nervous System / Q000737:chemistry / Q000378:metabolism*; D015180:Electrophoresis, Gel, Two-Dimensional; D000939:Epitopes / Q000134:cerebrospinal fluid / Q000276:immunology*; D005260:Female; D015151:Immunoblotting; D007150:Immunohistochemistry; D008969:Molecular Sequence Data; D009479:Neuropeptides / Q000134:cerebrospinal fluid* / Q000276:immunology / Q000378:metabolism; D009687:Nuclear Proteins; D011487:Protein Conformation; D016601:RNA-Binding Proteins; D000068103:Serine-Arginine Splicing Factors; D013552:Swine
D000906:Antibodies; D000939:Epitopes; D009479:Neuropeptides; D009687:Nuclear Proteins; D016601:RNA-Binding Proteins; C049628:antisecretory factor; D000068103:Serine-Arginine Splicing Factors