Translational profile of developing phellem cells in Arabidopsis thaliana roots.

The phellem is a specialized boundary tissue providing the first line of defense against abiotic and biotic stresses in organs undergoing secondary growth. Phellem cells undergo several differentiation steps, which include cell wall suberization, cell expansion, and programmed cell death. Yet, the molecular players acting particularly in phellem cell differentiation remain poorly described, particularly in the widely used model plant Arabidopsis thaliana. Using specific marker lines we followed the onset and progression of phellem differentiation in A. thaliana roots and further targeted the translatome of newly developed phellem cells using translating ribosome affinity purification followed by mRNA sequencing (TRAP-SEQ). We showed that phellem suberization is initiated early after phellogen (cork cambium) division. The specific translational landscape was organized in three main domains related to energy production, synthesis and transport of cell wall components, and response to stimulus. Novel players in phellem differentiation related to suberin monomer transport and assembly as well as novel transcription regulators were identified. This strategy provided an unprecedented resolution of the translatome of developing phellem cells, giving a detailed and specific view on the molecular mechanisms acting on cell differentiation in periderm tissues of the model plant Arabidopsis.

Pediatric Kidney Transplantation-Living or Deceased Donor?

BACKGROUND: Kidney transplantation is ideal for children and adolescents with chronic end-stage renal disease because it offers better growth, development, and quality of life. Donor choice is vitally important in this age group, given the long life expectancy of these patients. METHODS: A retrospective analysis of pediatric patients (<18 years) who underwent kidney transplantation from January 1999 to December/2018 was performed. Short- and long-term outcomes were compared between living and deceased donor transplants. RESULTS: We included 59 pediatric kidney transplant recipients, 12 from a living donor and 47 from a deceased donor. Thirty-six (61.0%) patients were boys, and 5 (8.5%) had a retransplant. There were no differences between groups on sex, race, and weight of the recipient and donor, as well as the age and the etiology of the recipient's primary disease. Most recipients received induction immunosuppression with basiliximab and maintenance with triple therapy, with no differences between groups. Living donor transplants were mostly pre-emptive (58.3% vs 4.3%, P < .001) and had fewer HLA mismatches (≤3: 90.9% vs 13.0%, P < .001), older donors (38.4 vs 24.3 years, P < .001) and shorter hospital stays (8.8 vs 14.1 days, P = .004). There were no statistically significant differences regarding medical-surgical complications and graft or patient survival. However, we found that at 13 years post-transplant 91.7% of the living donor grafts were functioning vs 72.3% of the deceased donor grafts. CONCLUSION: Our experience points out that a living donor graft in pediatric patients is associated with a higher probability of pre-emptive transplant, shorter hospital stay, greater HLA compatibility, and increased graft survival.

Spatiotemporal development of suberized barriers in cork oak taproots.

The longevity and high activity of the cork cambium (or phellogen) from Quercus suber L. (cork oak) are the cornerstones for the sustainable exploitation of a unique raw material. Cork oak is a symbolic model to study cork development and cell wall suberization, yet most genetic and molecular studies on these topics have targeted other model plants. In this study, we explored the potential of taproots as a model system to study phellem development and suberization in cork oak, thereby avoiding the time constraints imposed when studying whole plants. In roots, suberin deposition is found in mature endodermis cells during primary development and in phellem cells during secondary development. By investigating the spatiotemporal characteristics of both endodermis and phellem suberization in young seedling taproots, we demonstrated that secondary growth and phellogen activity are initiated very early in cork oak taproots (approx. 8 days after sowing). We further compared the transcriptomic profile of root segments undergoing primary (PD) and secondary development (SD) and identified multiple candidate genes with predicted roles in cell wall modifications, mainly lignification and suberization, in addition to several regulatory genes, particularly transcription factor- and hormone-related genes. Our results indicate that the molecular regulation of suberization and secondary development in cork oak roots is relatively conserved with other species. The provided morphological characterization creates new opportunities to allow a faster assessment of phellogen activity (as compared with studies using stem tissues) and to tackle fundamental questions regarding its regulation.

The Combined Effect of Heat and Osmotic Stress on Suberization of Arabidopsis Roots.

The simultaneous occurrence of heat stress and drought is becoming more regular as a consequence of climate change, causing extensive agricultural losses. The application of either heat or osmotic stress increase cell-wall suberization in different tissues, which may play a role in improving plant resilience. In this work, we studied how the suberization process is affected by the combination of drought and heat stress by following the expression of suberin biosynthesis genes, cell-wall suberization and the chemical composition in Arabidopsis roots. The Arabidopsis plants used in this study were at the onset of secondary root development. At this point, one can observe a developmental gradient in the main root, with primary development closer to the root tip and secondary development, confirmed by the suberized phellem, closer to the shoot. Remarkably, we found a differential response depending on the root zone. The combination of drought and heat stress increased cell wall suberization in main root segments undergoing secondary development and in lateral roots (LRs), while the main root zone, at primary development stage, was not particularly affected. We also found differences in the overall chemical composition of the cell walls in both root zones in response to combined stress. The data gathered showed that, under combined drought and heat stress, Arabidopsis roots undergo differential cell wall remodeling depending on developmental stage, with modifications in the biosynthesis and/or assembly of major cell wall components.

Enzymatic properties, evidence for in vivo expression, and intracellular localization of shewasin D, the pepsin homolog from Shewanella denitrificans.

The widespread presence of pepsin-like enzymes in eukaryotes together with their relevance in the control of multiple biological processes is reflected in the large number of studies published so far for this family of enzymes. By contrast, pepsin homologs from bacteria have only recently started to be characterized. The work with recombinant shewasin A from Shewanella amazonensis provided the first documentation of this activity in prokaryotes. Here we extend our studies to shewasin D, the pepsin homolog from Shewanella denitrificans, to gain further insight into this group of bacterial peptidases that likely represent ancestral versions of modern eukaryotic pepsin-like enzymes. We demonstrate that the enzymatic properties of recombinant shewasin D are strongly reminiscent of eukaryotic pepsin homologues. We determined the specificity preferences of both shewasin D and shewasin A using proteome-derived peptide libraries and observed remarkable similarities between both shewasins and eukaryotic pepsins, in particular with BACE-1, thereby confirming their phylogenetic proximity. Moreover, we provide first evidence of expression of active shewasin D in S. denitrificans cells, confirming its activity at acidic pH and inhibition by pepstatin. Finally, our results revealed an unprecedented localization for a family A1 member by demonstrating that native shewasin D accumulates preferentially in the cytoplasm.