Annexin II is a growth-regulated gene, whose expression is significantly increased in various human cancers. We examined annexin II expression in II human B-cell lymphoma cell lines and in normal B-cells. Wide variation was observed in the levels of annexin II in these cell lines. Annexin II overexpression was observed in 5 cell lines, while significantly reduced expression was observed in Raji, OMA-BL-1 and REH cell lines. Analysis of the annexin II gene, mRNA and protein in Raji and OMA-BL-1 cell lines indicated that annexin II gene was unaltered and that a low level of annexin II transcripts are produced in these cells. Down-regulation of annexin II expression was at the transcriptional level, and no reexpression of annexin II was observed after treatment of cells with demethylating agents. Thus methylation of the annexin II gene does not appear to be responsible for annexin II down-regulation. A slow migrating altered form of annexin II was detected in Raji and OMA-BL-1 cells, which was detected with the anti-chicken annexin II antiserum, but not with the anti-human annexin II antiserum. The slow migrating annexin II species was found to be sensitive to dephosphorylation by calf intestinal alkaline phosphatase, resulting in reduction of the size of the protein on SDS-polyacrylamide gels. The phosphorylated annexin II was also observed in nuclear extracts of human K562 and HeLa cells. Thus, Raji and OMA-BL-1 cells exclusively produce a phosphorylated form of annexin II, and phosphorylated annexin II may be important for cell survival and proliferation.

Annexins I and II have been proposed to participate in regulation of inflammation and cell proliferation. In the present study, we examined the expression of annexins I and II in various levels of the bovine respiratory tract and in cultured bovine bronchial epithelial cells. Immunoblot analysis of whole-tissue extracts revealed low-level expression of annexins I and II in bronchial mucosa mainstem to fourth generation. In contrast, high levels of annexins I and II (15- to 20-fold higher than in the bronchial mucosa) were seen in the distal lung parenchyma. Immunohistochemical staining of tracheobronchial tissue sections with anti-annexin I antibody revealed an uneven positive reaction of about 30 to 40% of the columnar differentiated ciliated and secretory cells, with no obvious positivity within the nondifferentiated basal epithelial cell layer. In contrast, anti-annexin II antibody reacted nearly uniformly within the basal cell layer, with no obvious decoration of the differentiated cell types. These results were confirmed by immunoblot analysis of annexin I and II in density fractionated populations of basal and secretory/ciliated epithelial cells. Both annexins I and II were expressed at constitutive levels in bovine bronchial epithelial cells grown in glucocorticoid- and serum-free medium, and dexamethasone increased expression of both proteins in a concentration-dependent manner. We conclude that annexins I and II are differentially expressed in the differentiated ciliated cells and undifferentiated basal cells.(ABSTRACT TRUNCATED AT 250 WORDS)

Primer recognition proteins (PRP) enable DNA polymerase alpha to utilize efficiently DNA substrates with low primer to template ratios. We have previously identified the protein-tyrosine kinase substrate annexin II, and the glycolytic enzyme 3-phosphoglycerate kinase as components of PRP. As a step towards elucidation of the role of PRP in the process of DNA replication, we have investigated the subcellular distribution and specific association of these proteins with the nuclear matrix in HeLa cells. Nuclear extracts prepared from HeLa cells in S phase contain the enzymatic activity of 3-phosphoglycerate kinase (PGK) and phospholipase A2 inhibitory activity of annexin II. Monomer annexin II is approximately equally distributed between the nuclear and cytoplasmic fractions, while a majority of PGK is in the cytoplasm. Immunoblot analyses reveal the presence of these two proteins in nuclei, specifically associated with the nuclear matrix. This is further confirmed by observation of the presence of annexin II and PGK in isolated nuclear matrices by immunoelectron microscopy. The phospholipase A2 inhibitory activity of annexin II colocalizes with the nuclear matrix-bound annexin II. A related protein, annexin I, is not detectable in the nuclear extracts and nuclear matrix. A slower-migrating (perhaps modified) form of annexin II is found to be associated with the nuclear matrix. Attempts to dissociate PGK and annexin II from the nuclear matrix with octyl-beta-glucoside, high salt or metal ion chelators were unsuccessful, suggesting that the interaction is very strong.

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