The mitogen-activated protein kinase pathway mediates estrogen neuroprotection after glutamate toxicity in primary cortical neurons.

Pharmacological and biochemical approaches were used to elucidate the involvement of growth factor signaling pathways mediating estrogen neuroprotection in primary cortical neurons after glutamate excitotoxicity. We addressed the activation of mitogen-activated protein kinase (MAPK) signaling pathways, which are activated by growth factors such as nerve growth factor (NGF). Inhibition of MAPK signaling with the MAPK kinase inhibitor PD98059 blocks both NGF and estrogen neuroprotection in these neurons. These results correlate with a rapid and sustained increase in MAPK activity within 30 min of estrogen exposure. The involvement of signaling molecules upstream from MAPK was also examined to determine whether activation of MAPK by estrogen is mediated by tyrosine kinase activity. Estrogen produces a rapid, transient activation of src-family tyrosine kinases and tyrosine phosphorylation of p21(ras)-guanine nucleotide activating protein. Effects of estrogen on neuroprotection, as well as rapid activation of tyrosine kinase and MAPK activity, are blocked by the anti-estrogen ICI 182,780. This provides evidence that activation of the MAPK pathway by estrogen participates in mediating neuroprotection via an estrogen receptor. These results describe a novel mechanism by which cytoplasmic actions of the estrogen receptor may activate the MAPK pathway, thus broadening the understanding of effects of estrogen in neurons.

Isoform-specific, calcium-regulated interaction of the synaptic vesicle proteins SV2 and synaptotagmin.

The identification and functional characterization of proteins localized to synaptic vesicles has contributed significantly to our understanding of neurotransmission. Studies of synaptic vesicle protein interactions have both led to the identification of novel synaptic proteins and suggested hypotheses of protein function. Synaptic vesicle protein 2 (SV2), is an integral membrane glycoprotein present in all synaptic vesicles. There are two characterized isoforms, SV2A and SV2B. Despite their homology to transporter proteins, the function of the SV2s remains unknown. In an effort to determine SV2 function and identify cofactors required for SV2 activity, we examined the protein interactions of SV2 using a combination of cross-linking, immunoprecipitation, and recombinant protein affinity chromatography. We report that SV2 is part of a large protein complex that contains the synaptic vesicle protein synaptotagmin. The interaction between SV2 and synaptotagmin is direct, specific to SV2A, and inhibited by calcium with an EC50 of approximately 10 microM. Interaction is mediated by the cytoplasmic amino terminus of SV2A and the C2B domain of synaptotagmin. Our observations suggest a regulatory relationship between these two proteins.

A germline substitution in the human MSH2 gene is associated with high-grade dysplasia and cancer in ulcerative colitis.

BACKGROUND & AIMS: The DNA mismatch repair gene human MSH2 shows a germline mutation in certain family members with hereditary nonpolyposis colorectal cancer. There is an increased risk of colorectal cancer in patients with ulcerative colitis (UC) with extensive disease of > 8 years' duration; however, specific constitutional predisposing genetic abnormalities have not yet been identified. METHODS: A germline human MSH2 abnormality was sought in patients with UC with high-grade dysplasia or carcinoma. RESULTS: After direct sequencing of exon 13 and flanking regions of human MSH2, a germline T to C substitution was shown at the -6 intronic splice acceptor site of exon 13. This substitution was found in 14 of 53 patients with UC with high-grade dysplasia or carcinoma (26%) compared with 4 of 36 high-risk patients with UC without dysplasia or cancer (11%) (P < or = 0.04) and in 7 of 80 healthy adult blood donors (9%) (P < or = 0.003). The patients with UC who had the substitution were three times more likely to develop neoplasia than patients with UC who did not carry it. CONCLUSIONS: An intronic splice-site substitution in the human MSH2 gene is present in the general population but may predispose to cancer in the setting of UC.

Deletion mapping of chromosome 8p in colorectal carcinoma and dysplasia arising in ulcerative colitis, prostatic carcinoma, and malignant fibrous histiocytomas.

Short tandem repeat polymorphism markers on the short arm of chromosome 8 were used to search for loss of heterozygosity (LOH) in colorectal carcinoma and dysplasia complicating ulcerative colitis, in prostatic carcinoma, and in malignant fibrous histiocytoma (MFH). Fifty percent of prostatic carcinomas (13/26), 44% of carcinomas or dysplasias arising in ulcerative colitis (7/16), and 30% (4/12) of MFH cases showed LOH for markers on 8p. Detailed mapping demonstrated variability in the size of the chromosomal region showing LOH; however, the data suggest a common 30-centimorgan region of LOH on chromosome 8p between the LPL locus and pter in colorectal and prostatic cancers. In addition, LOH was observed on 8p in both high-grade and low-grade dysplasia in ulcerative colitis, indicating that LOH on 8p may occur at an early stage of neoplastic development in this disorder. In contrast, MFH cases exhibited LOH for marker D8S87, which has been identified as being near the putative Werner's syndrome locus. These results suggest that a tumor suppressor gene, located on the distal portion of chromosome 8p, exists in common for prostatic and colorectal carcinomas, and a second tumor suppressor gene may exist linked to the Werner's syndrome locus.