Diversity of Saxicolous Lichens along an Aridity Gradient in Central México

Lichens are symbiotic organisms that comprise a fungus and a photosynthetic partner wich are recognized as a good indicator of climate change. However, our understanding of how aridity affects the diversity of saxicolous lichens in drylands is still limited. To evaluate the relationship between saxicolous lichen diversity and aridity in a central México dryland, a geographical transect was established of 100 km to build an aridity gradient in the semiarid zone of the State of Querétaro, Mexico, comprising ten sampling sites with a 10 km separation. Species richness, abundance and diversity of soil lichen species were recorded using two sampling methods: the quadrat-intercept and the line-intercept method, to compare their performance in assessing soil lichen diversity in drylands. The number of species and Shannon diversity of saxicolous lichens were higher at intermediate values of the aridity index (AI = 0.10–0.34). Quadrat intercept and point intercept methods gave quite similar results, which means that the selected method does not influence the results in a significant way. This study confirms the role of saxicolous lichens as climate change indicators and reveals the importance of the sampling method selection in the evaluation of different parameters of soil lichen diversity in drylands.     

No differential effects of divergent isocaloric supplements on signaling for muscle protein turnover during recovery from muscle-damaging eccentric exercise

Unaccustomed high-intensity eccentric exercise (ECC) can provoke muscle damage including several days of muscle force loss. Post-exercise dietary supplementation may provide a strategy to accelerate rate of force regain by affecting mechanisms related to muscle protein turnover. The aim of the current study was to investigate if protein signaling mechanisms involved in muscle protein turnover would be differentially affected by supplementation with either whey protein hydrolysate and carbohydrate (WPH+CHO) versus isocaloric carbohydrate (CHO) after muscle-damaging ECC. Twenty-four young healthy participants received either WPH+CHO (n = 12) or CHO supplements (n = 12) during post-exercise recovery from 150 maximal unilateral eccentric contractions. Prior to, at 3 h and at 24, 48, 96 and/or 168 h post-exercise, muscle strength, muscle soreness, and Akt-mTOR and FOXO signaling proteins, were measured in an ECC exercising leg and in the contralateral non-exercise control leg (CON). After ECC, muscle force decreased by 23–27 % at 24 h post-exercise, which was followed by gradual, although not full recovery at 168 h post-exercise, with no differences between supplement groups. Phosphorylation of mTOR, p70S6K and rpS6 increased and phosphorylation of FOXO1 and FOXO3 decreased in the ECC leg, with no differences between supplement groups. Phosphorylation changes were also observed for rpS6, FOXO1 and FOXO3a in the CON leg, suggesting occurrence of remote tissue effects. In conclusion, divergent dietary supplementation types did not produce differences in signaling for muscle turnover during recovery from muscle-damaging exercise.     

Proline dehydrogenase is a positive regulator of cell death in different kingdoms

Proline dehydrogenase (ProDH) catalyzes the flavin-dependent oxidation of Pro into Δ1-pyrroline-5-carboxylate (P5C). This is the first of the two enzymatic reactions that convert proline (Pro) into glutamic acid (Glu). The P5C thus produced is non-enzymatically transformed into glutamate semialdehyde (GSA), which acts as a substrate of P5C dehydrogenase (P5CDH) to generate Glu. Activation of ProDH can generate different effects depending on the behavior of other enzymes of this metabolism. Under different conditions it can generate toxic levels of P5C, alter the cellular redox homeostasis and even produce reactive oxygen species (ROS). Recent studies indicate that in Arabidopsis, the enzyme potentiates the oxidative burst and cell death associated to the Hypersensitive Responses (HR). Interestingly, activation of ProDH can also produce harmful effects in other organisms, suggesting that the enzyme may play a conserved role in the control of cell death.Key words: proline, proline dehydrogenase, cell death, hypersensitive response (HR), reactive oxygen species (ROS)     

Recombinant expression of ShPI-1A, a non-specific BPTI-Kunitz-type inhibitor, and its protection effect on proteolytic degradation of recombinant human miniproinsulin expressed in Pichia pastoris

Pichia pastoris is a highly successful system for the large-scale expression of heterologous proteins, with the added capability of performing most eukaryotic post-translational modifications. However, this system has one significant disadvantage - frequent proteolytic degradation by P. pastoris proteases of heterologously expressed proteins. Several methods have been proposed to address this problem, but none has proven fully effective. We tested the effectiveness of a broad specificity protease inhibitor to control proteolysis. A recombinant variant of the BPTI-Kunitz protease inhibitor ShPI-1 isolated from the sea anemone Stichodactyla helianthus, was expressed in P. pastoris. The recombinant inhibitor (rShPI-1A), containing four additional amino acids (EAEA) at the N-terminus, was folded similarly to the natural inhibitor, as assessed by circular dichroism. rShPI-1A had broad protease specificity, inhibiting serine, aspartic, and cysteine proteases similarly to the natural inhibitor. rShPI-1A protected a model protein, recombinant human miniproinsulin (rhMPI), from proteolytic degradation during expression in P. pastoris. The addition of purified rShPI-1A at the beginning of the induction phase significantly protected rhMPI from proteolysis in culture broth. The results suggest that a broad specificity protease inhibitor such as rShPI-1A can be used to improve the yield of recombinant proteins secreted from P. pastoris.     

Regulation of the membrane-localized prostaglandin E synthases mPGES-1 and mPGES-2 in cardiac myocytes and fibroblasts

The proinflammatory mediator cyclooxygenase (COX)-2 and its product PGE(2) are induced in the ischemic heart, contributing to inflammatory cell infiltration, fibroblast proliferation, and cardiac hypertrophy. PGE(2) synthesis coupled to COX-2 involves two membrane-localized PGE synthases, mPGES-1 and mPGES-2; however, it is not clear how these synthases are regulated in cardiac myocytes and fibroblasts. To study this, we used primary cultures of neonatal ventricular myocytes (VM) and fibroblasts (VF) treated with IL-1beta for 24 h. To test for involvement of MAPKs in IL-1beta regulation of mPGES-1 and-2, cells were pretreated with the pharmacological inhibitors of p42/44 MAPK, p38 MAPK, and c-Jun kinase (JNK). mRNA was analyzed by RT-PCR. Protein was analyzed by densitometry of Western blots. mPGES-1 was undetectable in untreated VF but induced by IL-1beta; inhibition of either p42/44 MAPK or JNK, but not p38 MAPK, was almost completely inhibitory. In VM, inhibition of the three MAPKs reduced IL-1beta-stimulated mPGES-1 protein by 70-90%. mPGES-2 was constitutively synthesized in both VM and VF and was not regulated by IL-1beta or MAPKs. Confocal microscopy revealed colocalization of both mPGES-1 and mPGES-2 with COX-2 in the perinuclear area of both VF and VM. Finally, PGE(2) production was higher in VM than VF. Our data show that 1) mPGES-1 is induced in both VF and VM, 2) regulation of mPGES-1 by MAPK family members is different in the two cell types, 3) mPGES-2 is constitutively synthesized in both VM and VF and is not regulated, and 4) mPGES-1 and mPGES-2 are colocalized with COX-2 in both cells. Thus differences in activity of mPGES-1 and COX-2 or coupling of COX-2 with mPGES-1 may contribute to differences in PGE(2) production by myocytes and fibroblasts.