Hemopressins and other hemoglobin‐derived peptides in mouse brain: comparison between brain, blood, and heart peptidome and regulation in Cpe^fat/fat mice

Abstract

J. Neurochem. (2010) 113, 871–880. Abstract Many hemoglobin‐derived peptides are present in mouse brain, and several of these have bioactive properties including the hemopressins, a related series of peptides that bind to cannabinoid CB1 receptors. Although hemoglobin is a major component of red blood cells, it is also present in neurons and glia. To examine whether the hemoglobin‐derived peptides in brain are similar to those present in blood and heart, we used a peptidomics approach involving mass spectrometry. Many hemoglobin‐derived peptides are found only in brain and not in blood, whereas all hemoglobin‐derived peptides found in heart were also seen in blood. Thus, it is likely that the majority of the hemoglobin‐derived peptides detected in brain are produced from brain hemoglobin and not erythrocytes. We also examined if the hemopressins and other major hemoglobin‐derived peptides were regulated in the Cpefat/fat mouse; previously these mice were reported to have elevated levels of several hemoglobin‐derived peptides. Many, but not all of the hemoglobin‐derived peptides were elevated in several brain regions of the Cpefat/fat mouse. Taken together, these findings suggest that the post‐translational processing of alpha and beta hemoglobin into the hemopressins, as well as other peptides, is up‐regulated in some but not all Cpefat/fat mouse brain regions.

Similar Intracellular Peptide Profile of TAP1/β2 Microglobulin Double-Knockout Mice and C57BL/6 Wild-Type Mice as Revealed by Peptidomic Analysis

“…In agreement with the original hypothesis that natural intracellular peptides have biological activity, the overexpression of the intracellular peptide metabolizing enzyme thimet oligopeptidase, which is known to alter the intracellular peptide profile in HEK293 cells (18), similarly changed angiotensin II and isoproterenol cell activation measured by the luciferase gene reporter assay (22). Taken together, these results suggest the exciting possibility that at least some of the naturally occurring intracellular peptides described here and elsewhere (19,20,36) may have biological activities similar to those previously described (22).…”

“…All of the 92 peptides identified in the present study have also been identified in previous peptidomic studies using brain extracts from mice with distinctive genetic backgrounds, such as the BALB/C (19) and C57BKS/J (36). Altogether, these data suggest that mouse neuronal cells have a constant pool of intracellular peptides that is not related to MHC-I antigens.…”

“…The possibility that intracellular peptides are not solely related to antigen presentation opens new perspectives on their function as novel, biologically active molecules with roles in cell function, health, and disease. One exciting possibility is that these intracellular peptides described here and elsewhere (19,20,36) could be related to cell signaling. One of the features of this new class of terminal fragment of the neurogranin peptide KGPGPGGPGG AGGARGGAGGGPSGD is the one that is greatly increased in β2m/ TAP1 −/− mice, although in these mice, the distribution of the peptides seems more disperse than in the C57BL/6 wild-type mice putatively bioactive molecules (intracellular peptides) is their dual location within and outside of cells, which gives them the freedom to potentially interfere with cell signaling both at the plasma membrane receptor and at the signal transduction pathway levels.…”

“…In the present study, a semi-quantitative peptidomic assay using stable isotopic labels and mass spectrometry that has been extensively characterized (18,19,35,36) was used to examine possible differences between the intracellular peptide profile of whole brain tissue from TAP1/β2m −/− mice and C57BL/6 wild-type mice. We chose to investigate the peptide profile of the whole brain instead of specific brain areas because our main goal was to identify possible differences in the peptide diversity of TAP1/β2m −/− mice compared with C57BL/6 wild-type mice rather than to quantitate differences related to specific brain regions.…”

Cytosolic Carboxypeptidase 1 Is Involved in Processing α- and β-Tubulin

“…After incubation for 72 h at 37°C, cells were collected, washed four times in 30 ml of PBS by centrifugation for 5 min at 800 ϫ g at 4°C, and peptides extracted and labeled as described (31,32). Briefly, extracted peptides were labeled with five different 4-trimethylammoniumbutyryl isotopic labels (31,33). Duplicate samples treated with CCP1 siRNA were labeled with the D3-and D9-4-trimethylammoniumbutyryl, and triplicate control samples were labeled with D0-, D6-, and D12-4-trimethylammoniumbutyryl tags.…”