Structure-function analysis of plant G-protein regulatory mechanisms identifies key Gα-RGS protein interactions.

Heterotrimeric GTP-binding protein alpha subunit (Gα) and its cognate regulator of G-protein signaling (RGS) protein transduce signals in eukaryotes spanning protists, amoeba, animals, fungi, and plants. The core catalytic mechanisms of the GTPase activity of Gα and the interaction interface with RGS for the acceleration of GTP hydrolysis seem to be conserved across these groups; however, the RGS gene is under low selective pressure in plants, resulting in its frequent loss. Our current understanding of the structural basis of Gα:RGS regulation in plants has been shaped by Arabidopsis Gα, (AtGPA1), which has a cognate RGS protein. To gain a comprehensive understanding of this regulation beyond Arabidopsis, we obtained the x-ray crystal structures of Oryza sativa Gα, which has no RGS, and Selaginella moellendorffi (a lycophyte) Gα that has low sequence similarity with AtGPA1 but has an RGS. We show that the three-dimensional structure, protein-protein interaction with RGS, and the dynamic features of these Gα are similar to AtGPA1 and metazoan Gα. Molecular dynamic simulation of the Gα-RGS interaction identifies the contacts established by specific residues of the switch regions of GTP-bound Gα, crucial for this interaction, but finds no significant difference due to specific amino acid substitutions. Together, our data provide valuable insights into the regulatory mechanisms of plant G-proteins but do not support the hypothesis of adaptive co-evolution of Gα:RGS proteins in plants.

Overexpression of a Plant-Specific Gγ Protein, AGG3, in the Model Monocot Setaria viridis Confers Tolerance to Heat Stress.

The vascular plant-specific, cysteine-rich type III Gγ proteins, which are integral components of the heterotrimeric G-protein complex, play crucial roles in regulating a multitude of plant processes, including those related to crop yield and responses to abiotic stresses. The presence of multiple copies of type III Gγ proteins in most plants and a propensity of the presence of specific truncated alleles in many cultivated crops present an ambiguous picture of their roles in modulating specific responses. AGG3 is a canonical type III Gγ protein of Arabidopsis, and its overexpression in additional model crops offers the opportunity to directly evaluate the effects of protein expression levels on plant phenotypes. We have shown that AGG3 overexpression in the monocot model Setaria viridis leads to an increase in seed yield. In this study, we have investigated the response of the S. viridis plants overexpressing AGG3 to heat stress (HS), one of the most important abiotic stresses affecting crops worldwide. We show that a short span of HS at a crucial developmental time point has a significant effect on plant yield in the later stages. We also show that plants with higher levels of AGG3 are more tolerant to HS. This is attributed to an altered regulation of stress-responsive genes and improved modulation of the photosynthetic efficiency during the stress. Overall, our results confirm that AGG3 plays a crucial role in regulating plant responses to unfavorable environmental conditions and may contribute positively to avoiding crop yield losses.

Low-cost homemade cocktails for enzymatic conversion of sugarcane and cassava bagasses.

The agricultural industries generate lignocellulosic wastes that can be modified by fungi to generate high value-added products. This work aimed to analyze the efficiency and the cost-effectiveness of the bioconversion of sugarcane and cassava bagasses using low-cost homemade enzymatic cocktails from Aspergillus niger LBM 134. Both bagasses were pretreated with a soft alkaline solution without any loss of polysaccharides. After the hydrolysis, a 28% of conversion to glucose and 42% to xylose were reached in the hydrolysis of sugarcane bagasse while an 80% of saccharification yield, in the hydrolysis of cassava bagasse using the homemade enzymes. Furthermore, a more disorganised surface and no starch granules were observed in the sugarcane and cassava bagasses, respectively. The bioethanol yield from sugarcane and casava bagasses was predicted to be 4.16 mg mL-1 and 2.57 mg mL-1, respectively. A comparison of the cost of the homemade and the commercial enzymes was carried out. Similar hydrolysis percentages were achieved employing any enzyme; however, it was 1000-2000 times less expensive using the homemade cocktails than using the commercial enzymes. Therefore, the cost of obtaining glucose from bagasses was most expensive when applying the commercial enzymes. Moreover, the hydrolysis of the cassava bagasse was most efficient with the homemade cocktails. The importance and novelty of this work lie in the similar performance and the lower cost of the homemade cocktails from the fungus A. niger LBM 134 compared with the commercial enzymes on the hydrolysis of the sugarcane and cassava bagasses.

DNA barcodes highlight genetic diversity patterns in rodents from lowland desert and Andean areas in Argentina.

Rodents are an important component of South America fauna. Their high diversity has motivated researchers to continually review their taxonomy, genetic diversity, species limits, and phylogenetic relationships. Here, we applied DNA-barcodes for assessing the taxonomic and genetic diversity in the two major lineages of South American rodents: caviomorphs and sigmodontines. We analysed 335 COI barcodes in 34 morphologically determined species from 39 localities along central Andes and arid lands of Argentina. Neighbour-joining and maximum likelihood reconstruction provided clear separation between species. The Barcode Index number and Bayesian Poisson tree processes were used to confirm concordance between sequence clusters and species designations by taxonomy. We found deep divergence within the Phyllotis xanthopygus species complex, with distances up to 13.0% between geographically separated lineages. Minor divergences (3.30% and 2.52%) were found within Abrothrix hirta, and Tympanoctomys barrerae, respectively, with differentiation in their genetic lineages. Also, we documented geographically separated clusters for Akodon spegazzinii and A. oenos with up to 2.3% divergence, but clustering methods failed to distinguish them as different species. Sequence results show a clear barcode gap with a mean intraspecific divergence (0.56%) versus a minimum nearest-neighbour distance averaging (10.1%). Distances between congeneric species varied from 4.1 to 14%, with the exception of two related forms within Euneomys and the sister species Akodon spegazzinii and A. oenos. This study constitutes a substantial contribution to the global barcode reference library. It provides insights into the complex phylogeographic patterns and speciation scenarios in rodents, while highlighting areas that require in-depth taxonomic and integrative research.

High resolution crystal structure of NaTrxh from Nicotiana alata and its interaction with the S-RNase.

Thioredoxins are regulatory proteins that reduce disulfide bonds on target proteins. NaTrxh, which belongs to the plant thioredoxin family h subgroup 2, interacts and reduces the S-RNase enhancing its ribonuclease activity seven-fold, resulting an essential protein for pollen rejection inNicotiana.Here, the crystal structure of NaTrxh at 1.7 Å by X-ray diffraction is reported. NaTrxh conserves the typical fold observed in other thioredoxins from prokaryotes and eukaryotes, but it contains extensions towards both N- and C-termini.The NaTrxh N-terminal extension participates in the reduction of S-RNase, and in the structure reported here, this is orientated towards the reactive site. The interaction between SF11-RNase and the NaTrxh N-terminal was simulated and the short-lived complex observed lasted for a tenth of ns. Moreover, we identified certain amino acids as SF11-RNase-E155 and NaTrxh-M104 as good candidates to contribute to the stability of the complex. Furthermore, we simulated the reduction of the C153-C186 SF11-RNase disulfide bond and observed subtle changes that affect the entire core, which might explain the increase in the ribonuclease activity of S-RNase when it is reduced by NaTrxh.