Clypeatula cooperensis gen. n., sp. n., a new freshwater sponge (Porifera, Spongillidae) from the Rocky Mountains of Montana, USA.

A new genus and species of freshwater sponge, Clypeatula cooperensis, collected from three lakes in the Northern Rocky Mountains of Montana, USA, are described. The sponge grows as a hard, disc-shaped encrustation on the undersides of rocks and logs. It lacks microscleres and has amphioxeal megascleres that often show a slight midregion bulb and are usually covered with short, conical spines except at their tips. The sponge is also non-gemmulating, overwintering in a regressed state in which choanocyte chambers are reduced in number. Phylogenetic analyses of complete 18S rDNA sequences of C. cooperensis, Ephydatia muelleri, Spongilla lacustris and Eunapius fragilis suggest that C. cooperensis is more closely related to Ephydatia muelleri than to Spongilla lacustris or Eunapius fragilis. Our data, nonetheless, do not rule out the possibility that C. cooperensis is more closely related to the non-gemmulating sponges of Lake Baikal (Russia) than it is to Ephydatia muelleri. These phylogenetic analyses support the erection of a new genus, the monophyly of freshwater sponges belonging to the families Spongillidae and Lubomirskiidae, and the monophyly of demosponges.

Biochemical and immunochemical determination of (Na+ + K+)-ATPase content in subcellular fractions prepared from the avian salt gland: evidence for enzyme heterogeneity.

Subcellular membrane fractions were prepared from the salt glands of osmotically-stressed ducklings. Two fractions were characterized biochemically with respect to (Na+ + K+)-ATPase, alkaline phosphodiesterase I, succinate dehydrogenase, esterase, and galactosyltransferase activities and immunochemically with respect to (Na+ + K+)-ATPase. The ratios of the estimates of the (Na+ + K+)-ATPase contents obtained biochemically and immunochemically from the two fractions differed by more than 2 X. The results are consistent with the presence of at least two molecular species of (Na+ + K+)-ATPase, unevenly distributed between the two fractions.

Plasma membrane biogenesis in the avian salt gland: a biochemical and quantitative electron microscopic autoradiographic study.

The avian salt gland provides an ideal system for the study of plasma membrane (PM) biogenesis. Feeding ducklings 1% sodium chloride (salt stress) induces the secretory cells of the gland to synthesize large amounts of PM, which forms an extensive basolateral PM domain after 7-9 days of treatment. In the present study, the initial biosynthetic events following salt stress were investigated. In vivo studies using 3H-uridine indicated that increased rates of RNA synthesis could be detected by 2 hr after the beginning of salt stress and continued through at least 12 hr. Under in vitro conditions, increased rates of protein and glycoprotein synthesis (as monitored by 3H-leucine and 3H-fucose incorporation, respectively) were also detected after 2 hr and continued through 7-9 days. Increased levels of Na,K-ATPase, a specific secretory cell PM marker, were detected after 8 hr of treatment as monitored by specific activity and 3H-ouabain binding. Sodium dodecyl sulfate-polyacrylamide slab gel electrophoresis coupled with fluorography indicated that both 3H-leucine and 3H-fucose were incorporated into partially purified preparations of Na,K-ATPase isolated after 12 hr. Light microscopic autoradiographic analysis of pulse-chase experiments indicated that in secretory cells of 12-hr salt-stressed glands, 3H-leucine- and 3H-fucose-labelled products reached the cell periphery by 1-2 hr after the initial pulse. The incorporation of both tritiated precursors was predominantly associated with the secretory cells. Quantitative electron microscopic autoradiography indicated that 3H-leucine is initially taken up by elements of the rough endoplasmic reticulum (RER) and cytoplasm (5 min postpulse), subsequently transported to and concentrated within components of the Golgi apparatus (10 min of chase), and ultimately incorporated into all domains of the plasma membrane of secretory cells by 1-2 hr of chase. The data is consistent with a flow of newly synthesized membrane components from RER to Golgi to plasma membrane and is analogous to the pattern previously found for the synthesis and processing of PM proteins in a wide variety of cell types.

Partial immunochemical identity between (Na+ + K+)-ATPase and a membrane component extractable with chloroform: methanol.

An IgG fraction prepared from an antiserum against a holoenzyme preparation of (Na+ + K+)-ATPase precipitated a single antigen when samples of holoenzyme were subjected to crossed immunoelectrophoresis but precipitated an additional, immunochemically-related antigen when a plasma membrane-enriched fraction was subjected to crossed immunoelectrophoresis under the same conditions. The immunochemically-related antigen could be extracted from the plasma membrane fraction with CHCl3:CH3OH.