Recently we cloned and described ERp29, a novel 29-kDa endoplasmic reticulum (ER) protein that is widely expressed in rat tissues. Here we report our original isolation of ERp29 from dental enamel cells, and the comprehensive sequence analysis that correlated ERp29 with its cognate cDNA, both in enamel cells and liver. Fractionation of enamel cells using a new freeze±thaw procedure showed that ERp29 partitioned with known reticuloplasmins, and not with soluble proteins from mitochondria or cytosol. The absence of ERp29 in secreted enamel matrix indicated that the C-terminal tetrapeptide (KEEL motif ) confers effective ER-retention in enamel cells. ERp29 behaved as a single species (< 40 kDa) during size-exclusion chromatography of liver reticuloplasm, suggesting that most ERp29 is not stably associated with other proteins. Immunoblot analysis showed that ERp29 was up-regulated during enamel secretion and expressed most highly in secretory tissues, indicative of a role in secretory-protein synthesis. Unlike other reticuloplasmins, ERp29 was down-regulated during enamel mineralization and thereby dissociated from a calcium-handling role. Tissue-specific variations in ERp29 molecular abundance were revealed by quantification of reticuloplasmin mole ratios. In conclusion: (a) ERp29 is a novel reticuloplasmin of general functional importance; (b) a unique role in protein processing is implicit from the distinctive expression patterns and molecular structure; (c) ERp29 is primarily involved in normal protein secretory events, not the ER stress response; (d) a major role is likely in tissues where ERp29 was equimolar with established molecular chaperones and foldases. This study implicates ERp29 as a new member of the ER protein-processing machinery, and identifies tissues where the physiological role of ERp29 is most likely to be clearly manifested.

Serological and PCR techniques failed to detect Tomato spotted wilt virus (TSWV) infecting stunted tomato plants exhibiting leaf curling, purpling and crumpling. In contrast, Mass Spectrometry rapidly identified peptides generated from a~28kD protein cut from PAGE gels as TSWV N-protein within 5 days. The results indicate that the incidence of TSWV may be underestimated in the field. Cocksfoot mild mosaic virus (CMMV) was identified infecting Bromus diandrus using serological techniques. Peptides from a~25kD protein failed to identify with any protein on GenBank until a CMMV sequence was published. The use of Mass Spectrometry as an adjunct for virus detection and identification is briefly discussed.