Brain serine proteases are implicated in developmental processes, synaptic plasticity, and in disorders including Alzheimer's disease. The spectrum of the major enzymes expressed in brain has not been established previously. We now present a systematic study of the serine proteases expressed in adult rat and mouse hippocampus. Using a combination of techniques including polymerase chain reaction amplification and Northern blotting we show that tissue-type plasminogen activator (t-PA) is the major species represented. Unexpectedly, the next most abundant species were RNK-Met-1, a lymphocyte protease not reported previously in brain, and two new family members, BSP1 (brain serine protease 1) and BSP2. We report full-length sequences of the two new proteases; homologies indicate that these are of tryptic specificity. Although BSP2 is expressed in several brain regions, BSP1 expression is strikingly restricted to hippocampus. Other enzymes represented, but at lower levels, included elastase IV, proteinase 3, complement C2, chymotrypsin B, chymotrypsin-like protein, and Hageman factor. Although thrombin and urokinase-type plasminogen activator were not detected in the primary screen, low level expression was confirmed using specific polymerase chain reaction primers. In contrast, and despite robust expression of t-PA, the usual t-PA substrate plasminogen was not expressed at detectable levels.

hct-1 (hippocampal transcript) was detected in a differential screen of a rat hippocampal cDNA library. Expression of hct-1 was enriched in the formation but was also detected in rat liver and kidney, though at much lower levels; expression was barely detectable in testis, ovary, and adrenal. In liver, unlike brain, expression was sexually dimorphic; hepatic expression was greatly reduced in female rats. In mouse, brain expression was widespread, with the highest levels being detected in corpus callosum; only low levels were detected in liver. Sequence analysis of rat and mouse hct-1 cDNAs revealed extensive homologies with cytochrome P450s (CYPs), a diverse family of heme-binding monooxygenases that metabolize a range of substrates including steroids, fatty acids, and xenobiotics. Among the CYPs, hct-1 is most similar (39% at the amino acid sequence) to cholesterol 7␣-hydroxylase (CYP7) and contains a postulated steroidogenic domain present in other steroidmetabolizing CYPs but clearly represents a type of CYP not previously reported. Genomic Southern analysis suggests that a single gene corresponding to hct-1 is present in mouse, rat, and human. hct-1 is unusual in that, unlike all other CYPs described, the primary site of expression is in the brain. Similarity to CYP7 and other steroid-metabolizing CYPs may argue that hct-1 (CYP7B) plays a role in steroid metabolism in brain, notable because of the documented ability of brain-derived steroids (neurosteroids) to modulate cognitive function in vivo.Cytochromes P450, a diverse group of heme-containing monooxygenases (termed CYPs) 1 (for nomenclature see Nelson et al. (1)), catalyze a variety of oxidative conversions, notably of steroids but also of fatty acids and xenobiotics. Though most abundantly expressed in the testis, ovary, placenta, adrenal, and liver, the brain is a further site of CYP expression (2-6).Several CYP activities or mRNAs have been reported in the nervous system, predominantly of types metabolizing fatty acids and xenobiotics (subclasses CYP2C, -2D, -2E, and -4 (6, 7)). However, primary rat brain-derived glial cells can synthesize pregnenolone and progesterone in vitro (8). Mellon and Deschepper (9) provided molecular evidence for the presence, in brain, of key steroidogenic enzymes CYP11A1 (scc) and CYP11B1 (11␤) but failed to detect CYP17 (c17) or CYP11B2 (AS). Though CYP21A1 (c21) activity is reported to be present in brain (10) authentic CYP21A1 transcripts were not detected (11).Interest in brain steroid metabolism has been fueled by the finding that adrenal-and brain-derived steroids (neurosteroids) can modulate cognitive function and synaptic plasticity (reviewed in Refs. 12-17). For instance, pregnenolone and steroids derived from it are reported to have memory-enhancing effects in mice (18,19). However, the full spectrum of brain CYPs and the biological roles of their metabolites in vivo have not been established.To investigate such regulation of brain function our studies have focused on the hippocampus, a brain region important ...

Mice harboring random gene‐trap insertions of a lacZ (β‐galactosidase)‐neomycin resistance fusion cassette (β‐geo) were analyzed for expression in the hippocampus. In 4 of 15 lines reporter gene activity was observed in the hippocampal formation. In the obn line, enzyme activity was detected in the CA1–3 hippocampal subfields, in hpk expression was restricted to CA1, but in both lines reporter activity was also present in other brain regions. In the third line, kin, reporter activity was robustly expressed throughout the stratum pyrimidale of CA1–3, with only low‐level expression elsewhere. The final line (glnC) displayed ubiquitous expression of the reporter and was not analyzed further. Fusion transcripts for the first three lines were characterized; all encode polypeptides with features of membrane‐associated signalling proteins. The obn fusion identified a human cDNA (B2–1) encoding a pleckstrin homology (PH) domain, while hpk sequences matched the Epstein‐Barr Virus (EBV) inducible G‐protein coupled receptor, EBI‐1. kin identified an alternative form of the abl‐related nonreceptor tyrosine kinase c‐arg. Electrophysiological studies on mice homozygous for the insertions revealed normal synaptic transmission, paired pulse facilitation and paired‐pulse depression at Schaffer collateral‐commissural CA1 synapses, and normal long‐term potentiation (LTP) in obn and kin. hpk mice displayed an increase in hippocampal CA1 long‐term potentiation (LTP), suggesting a role for this receptor in synaptic plasticity. Hippocampus 1998;8:444–457. © 1998 Wiley‐Liss, Inc.

No abstract

No abstract