Proteasome activity and the post-translational control of sucrose synthase stability in maize leaves.

The serine-170 (S170) calcium-dependent protein kinase phosphorylation site of maize (Zea mays L.) sucrose synthase (SUS) (EC 2.4.1.13) has been implicated in the post-translational regulation of SUS protein stability. To clarify the proteolytic process and the role of phosphorylation, SUS degradation and proteasome activities were studied in the maize leaf elongation zone. Size-exclusion chromatography resolved two peaks of proteasome-like proteolytic activity. The large molecular mass ( approximately 1350 kDa) peak required Mg(2+) and ATP for maximal activity and was inhibited by the proteasome inhibitors MG132 and NLVS. Anion-exchange chromatography resolved a similar proteolytic activity that was activated by ATP, characteristics that are consistent with those of a 26S-proteasome. Appropriately, immunoblotting revealed the presence of a 26S-proteasome subunit and highly ubiquitinated proteins within the active fractions eluted from both columns. The smaller molecular mass ( approximately 600 kDa) peak represented only 40% of the total proteasome-like activity and is likely a maize 20S-proteasome as it was activated in vitro by low levels of sodium dodecyl sulfate (SDS). S170 phosphorylated SUS (pS170-SUS) was detected as both high molecular mass (HMM) forms and proteolytic fragments that co-eluted with 26S-proteasome activities on both size-exclusion and anion-exchange columns. Conditions that maintained maximal 26S-proteasome activity reduced the amounts of pS170-SUS recovered. In vitro, the 26S-proteasome degraded SUS and proteasome-specific inhibitors reduced SUS proteolysis. HMM-SUS conjugates were produced in vitro and immunoprecipitations suggested that some SUS might be ubiquitinated in vivo. The results suggest that S170 phosphorylation promotes the formation of HMM, ubiquitin-SUS conjugates that can be targeted for 26S-proteasome-dependent degradation.     

Brief debrisoquin administration to assess central dopaminergic function in children

Central dopaminergic (DA) function in children was assessed by monitoring plasma-free homovanillic acid (pHVA) levels after brief (18 hour) administration with debrisoquin sulfate, a peripherally active antihypertensive agent that blocks peripheral, but not central, HVA production. Brief debrisoquin administration resulted in marked reductions in pHVA in each of six patients studied. In five of the six patients, post-debrisoquin pHVA levels remained relatively stable over the six-hour period of observation. No significant cardiovascular or behavioral side effects of debrisoquin were observed. The brief debrisoquin administration method appears to be a safe, simple, and potentially valid peripheral technique for evaluating aspects of central dopaminergic function in children with neuropsychiatric disorders. Additional work is needed to further establish this method's validity and reliability.     

The role of lysyl oxidase and collagen crosslinking during sea urchin development

Lysyl oxidase, the only enzyme involved in collagen crosslinking, is shown to be present in embryos of the sea urchin Strongylocentrotus purpuratus. The enzyme specific activity increases over six-fold during development, showing the greatest rise during gastrulation and prism larva formation. The enzyme is inhibited by the specific inhibitor, beta-aminoproprionitrile (BAPN). Continuous BAPN treatment of S. purpuratus and Lytechinus pictus embryos from late cleavage stages onward increases the amount of noncrosslinked collagen present in prism larvae. When BAPN is added at the 128- or 256-cell stage it causes developmental arrest at the mesenchyme blastula stage. Embryos can be maintained in the arrested state for at least 96 h and will resume normal development and morphogenesis following BAPN removal. If BAPN is added after the mesenchyme blastula stage, it has little adverse effect on development; consequently nonspecific toxic effects of the drug are unlikely. The results suggest that lysyl oxidase and collagen crosslinking play a vital role in primary mesenchyme migration, gastrulation, and morphogenesis during sea urchin development and indicate that BAPN may be very useful in studying the extracellular matrix-cell interactions at the cellular and molecular level.     

Cloprostenol inouced luteal regression in the beef cow. I. Ovarian response and endocrine changes

Forty-two cycling, multiparous beef cows (percentage-Brahman) were given two injections of 500 ug Cloprostenol (CLP) 11 days apart. Cows were randomly allocated to be ovariectomized at 0, 12, 24, 36, 48, 60 or 72 hr after the second CLP injection. Mean CL weight declined within 36 hr after CLP. Mean concentration of P4 in luteal tissue increased between o and 60 hr, while mean P4 content per CL declined by 12 hr after CLP. There was a precipitous decrease in mean serum P4 by 12 hr following CLP injection. Serum E2 was elevated until 24 hr and then declined through 72 hr after CLP. Follicular T concentration increased from 0 to 48 hr and then decreased by 60 hr. We conclude that CLP caused rapid diminution of luteal function which was accompanied by a reduction in P4 content but not in P4 concentration. Futhermore, the concentration of E2 in large follicles decreased by 72 hr post-CLP which is consistent with an alteration of the steroidogenic pathway in the periovulatory follicle.     

Range position and climate sensitivity: The structure of among‐population demographic responses to climatic variation

Species’ distributions will respond to climate change based on the relationship between local demographic processes and climate and how this relationship varies based on range position. A rarely tested demographic prediction is that populations at the extremes of a species’ climate envelope (e.g., populations in areas with the highest mean annual temperature) will be most sensitive to local shifts in climate (i.e., warming). We tested this prediction using a dynamic species distribution model linking demographic rates to variation in temperature and precipitation for wood frogs (Lithobates sylvaticus) in North America. Using long‐term monitoring data from 746 populations in 27 study areas, we determined how climatic variation affected population growth rates and how these relationships varied with respect to long‐term climate. Some models supported the predicted pattern, with negative effects of extreme summer temperatures in hotter areas and positive effects on recruitment for summer water availability in drier areas. We also found evidence of interacting temperature and precipitation influencing population size, such as extreme heat having less of a negative effect in wetter areas. Other results were contrary to predictions, such as positive effects of summer water availability in wetter parts of the range and positive responses to winter warming especially in milder areas. In general, we found wood frogs were more sensitive to changes in temperature or temperature interacting with precipitation than to changes in precipitation alone. Our results suggest that sensitivity to changes in climate cannot be predicted simply by knowing locations within the species’ climate envelope. Many climate processes did not affect population growth rates in the predicted direction based on range position. Processes such as species‐interactions, local adaptation, and interactions with the physical landscape likely affect the responses we observed. Our work highlights the need to measure demographic responses to changing climate.