The small iron-deficiency-induced protein OLIVIA and its relation to the bHLH transcription factor POPEYE.

Iron (Fe) is a crucial micronutrient needed in many metabolic processes. To balance needs and potential toxicity, plants control the amount of Fe they take up and allocate to leaves and seeds during their development. One important regulator of this process is POPEYE (PYE). PYE is a Fe deficiency-induced key bHLH transcription factor (TF) for allocation of internal Fe in plants. In the absence of PYE, there is altered Fe translocation and plants develop a leaf chlorosis. NICOTIANAMINE SYNTHASE4 (NAS4), FERRIC-REDUCTION OXIDASE3 (FRO3), and ZINC-INDUCED FACILITATOR1 (ZIF1) genes are expressed at higher level in pye-1 indicating that PYE represses these genes. PYE activity is controlled in a yet unknown manner. Here, we show that a small Fe deficiency-induced protein OLIVIA (OLV) can interact with PYE. OLV has a conserved C-terminal motif, that we named TGIYY. Through deletion mapping, we pinpointed that OLV TGIYY and several regions of PYE can be involved in the protein interaction. An OLV overexpressing (OX) mutant line exhibited an enhanced NAS4 gene expression. This was a mild Fe deficiency response phenotype that was related to PYE function. Leaf rosettes of olv mutants remained smaller than those of wild type, indicating that OLV promotes plant growth. Taken together, our study identified a small protein OLV as a candidate that may connect aspects of Fe homeostasis with regulation of leaf growth.

Impact of Bronchoscopic Lung Volume Reduction with Valves on the Pulmonary Gas Exchange.

Introduction: Bronchoscopic lung volume reduction (BLVR) with endobronchial valves has been shown to be a safe and effective treatment for patients with severe lung emphysema. Previous studies have reported a benefit in pulmonary function, exercise capacity, and quality of life after BLVR-treatment. The effect of BLVR with valves on the pulmonary gas exchange and its association with clinical outcomes has not been analyzed to date. The primary goal of this study was to investigate the impact of BLVR on the pulmonary gas exchange and the impact of the target lobe selection in patients with discordant target lobes in high-resolution computed tomography (HRCT) scan and perfusion scan on the pulmonary gas exchange and clinical outcomes. Methods: In this single-center study, we retrospectively analyzed pulmonary function tests, 6-min-walk-tests, HRCT scans, perfusion scans, and blood gas analyses in 77 patients over the course of 6 months following BLVR treatment. Results: We observed that complete lobar occlusion with bronchoscopic valves leads to a transient impairment of pulmonary gas exchange. Despite this, an overall positive clinical outcome could be shown in patients treated with endobronchial valves. If the target lobe selection based on HRCT and perfusion scans is discrepant, a selection based on the HRCT scan tends to be associated with a better outcome than a selection based on the perfusion scan. Conclusions: Complete lobar occlusion with bronchoscopic valves leads to a transient impairment of pulmonary gas exchange but nevertheless results in an overall positive clinical outcome.

Survival outcomes in a prospective randomized multicenter Phase III trial comparing patients undergoing anatomical segmentectomy versus standard lobectomy for non-small cell lung cancer up to 2 cm.

OBJECTIVES: The oncological equivalence of anatomical segmentectomy for early stage non-small cell lung cancer (NSCLC) is still controversial. Primary aim of this study was survival outcomes in combination with improved quality of life after segmentectomy compared with lobectomy in patients with pathological stage Ia NSCLC (up to 2 cm, 7th edition) MATERIALS AND METHODS: We conducted a prospective, randomized, multicenter phase III trial to confirm the non-inferiority of segmentectomy to lobectomy in regard to prognosis (trial No. DRKS00004897). Patients were randomized to undergo either segmentectomy or lobectomy and followed up for 5-years survival and tumor recurrence. The 5-year hazard ratio comparing lobectomy with segmentectomy was required to remain above 0.5. RESULTS: Between October 2013 and June 2016, 108 patients with verified or suspected NSCLC up to 2 cm diameter were enrolled; 54 were assigned to lobectomy and 54 (1 drop-out) to segmentectomy. In-hospital and 90 days mortality was 0% in both groups. Overall survival at 5 years was 86.52% in the lobectomy compared to 78.21% in the segmentectomy group (HR = 0.61, (95% CI 0.23-1.66), p-value of non-inferiority test, p-ni = 0.687). Disease free survival was 77.29% for the lobectomy and 77.96% for the segmentectomy patients (HR = 1.50, (95% CI 0.60-3.76), p-ni = 0.019). At a median follow-up of 5 years, no differences were noted in either the locoregional or distant recurrent disease in both groups (9.4% vs 7.4%, p-ni = 0.506). CONCLUSION: Overall survival, locoregional and distant recurrences was not significantly difference for patients undergoing either segmentectomy or lobectomy for stage Ia NSCLC. The targeted non-inferiority of segmentectomy to lobectomy could not be proven for primary endpoint overall survival, but was significant for the secondary endpoint of disease free survival.

The Iron Deficiency-Regulated Small Protein Effector FEP3/IRON MAN1 Modulates Interaction of BRUTUS-LIKE1 With bHLH Subgroup IVc and POPEYE Transcription Factors.

In light of climate change and human population growth one of the most challenging tasks is to generate plants that are Fe-efficient, resilient to low Fe supply and Fe-biofortified. For such endeavors, it is crucial to understand the regulation of Fe acquisition and allocation in plants. One open question is how identified Fe-regulatory proteins comprising positive and negative regulators act together to steer Fe homeostasis. bHLH transcription factors (TFs) belonging to the subgroups IVb and IVc can initiate a bHLH cascade controlling the -Fe response in roots. In Arabidopsis thaliana, the -Fe-induced genes are sub-divided into several gene co-expression clusters controlled by different sets of TFs. Some of the co-expressed genes encode regulatory E3 ligase proteins BRUTUS (BTS)/BTS-LIKE (BTSL) and small proteins belonging to the group of FE UPTAKE-INDUCING PEPTIDE/IRON MAN (FEP/IMA). Recently, it was described that FEP1/IMA3 and FEP3/IMA1 proteins inhibit the repression of bHLH factors by BTS. We had postulated that -Fe-regulated co-expression clusters provide new information about regulatory protein interaction complexes. Here, we report a targeted yeast two-hybrid screen among 23 proteins of the -Fe response. This identified a novel protein interactome involving another E3 ligase, namely BTSL1, basic helix-loop-helix (bHLH) protein POPEYE (PYE) and transcription factors of the subgroup IVc as well as FEP3/IMA1. Because of the difficulty in stable BTSL1 protein expression in plant cells, we used a yeast two hybrid-based deletion mapping, homology modeling and molecular docking, to pinpoint interaction sites in BTSL1 and FEP3/IMA1. bHLH IVc TFs have similar residues at their C-terminus as FEP3/IMA1 interacting sites. FEP3/IMA1 attenuated interaction of BTSL1 and bHLH proteins in a yeast three-hybrid assay, in line with physiological data pointing to enhanced Fe acquisition and allocation in FEP3/IMA1 overexpression and btsl1 btsl2 mutant plants. Hence, exploiting -Fe-induced gene co-expression networks identified FEP3/IMA1 as a small effector protein that binds and inhibits the BTSL1 complex with PYE and bHLH subgroup IVc proteins. Structural analysis resolved interaction sites. This information helps improving models of Fe regulation and identifying novel targets for breeding of Fe-efficient crops.

Significantly favourable outcome for patients with non-small-cell lung cancer stage IIIA/IIIB and single-station persistent N2 (skip or additionally N1) disease after multimodality treatment.

OBJECTIVES: Persistent lymph nodes infiltration after neoadjuvant treatment remains a controversial topic in the treatment of stage III non-small-cell lung cancer (NSCLC). The aim of this study is to identify subgroups with persistent N2 disease, who could experience survival benefit from the addition of surgery. METHODS: A retrospective mono-institutional study was conducted to analyse all patients with a final histopathology of NSCLC and persistent mediastinal disease after induction chemotherapy or chemoradiotherapy and surgery from January 1998 to June 2015. RESULTS: A total of 145 patients (93 men, 52 women) fulfilled the inclusion criteria. The median age was 60 years (range 38-78). A total of 82 (56.5%) patients received a lobectomy, 48 (33.1%) a pneumonectomy, 11 (7.6%) a bilobectomy and 4 (2.6%) an anatomical segmentectomy; 128 (88.3%) were completely resected (R0). Operative mortality was 2.6% (4 patients), and morbidity was 35.2% (51 patients). Overall survival at 5 years was 47.3% (n = 19) for single N2 (skip), 30.2% (n = 16) for single N2 and N1 lymph nodes and under 5% (n = 1) for multiple mediastinal stations disease. Overall survival at 5 years after lobectomy/bilobectomy was not statistically different than after pneumonectomy (33.5% vs 20.5%, P = 0.082). Disease-free survival at 5 years was 30.6% (n = 6) for ypN2a1, 23.4% (n = 7) for ypN2a2 and under 5% (n = 1) for ypN2b status. CONCLUSIONS: Lobectomy or bilobectomy has to be taken into account as a potentially curative option with promising long-term results for patients after induction treatment and persistent single-station N2 involvement (skip or additionally N1 status). TRIAL REGISTRY NUMBER: 14-6138-BO.

Putative cis-Regulatory Elements Predict Iron Deficiency Responses in Arabidopsis Roots.

Plant iron deficiency (-Fe) activates a complex regulatory network that coordinates root Fe uptake and distribution to sink tissues. In Arabidopsis (Arabidopsis thaliana), FER-LIKE FE DEFICIENCY-INDUCED TRANSCRIPTION FACTOR (FIT), a basic helix-loop-helix (bHLH) transcription factor (TF), regulates root Fe acquisition genes. Many other -Fe-induced genes are FIT independent, and instead regulated by other bHLH TFs and by yet unknown TFs. The cis-regulatory code, that is, the cis-regulatory elements (CREs) and their combinations that regulate plant -Fe-responses, remains largely elusive. Using Arabidopsis root transcriptome data and coexpression clustering, we identified over 100 putative CREs (pCREs) that predicted -Fe-induced gene expression in computational models. To assess pCRE properties and possible functions, we used large-scale in vitro TF binding data, positional bias, and evolutionary conservation. As one example, our approach uncovered pCREs resembling IDE1 (iron deficiency-responsive element 1), a known grass -Fe response CRE. Arabidopsis IDE1-likes were associated with FIT-dependent gene expression, more specifically with biosynthesis of Fe-chelating compounds. Thus, IDE1 seems to be conserved in grass and nongrass species. Our pCREs matched among others in vitro binding sites of B3, NAC, bZIP, and TCP TFs, which might be regulators of -Fe responses. Altogether, our findings provide a comprehensive source of cis-regulatory information for -Fe-responsive genes that advance our mechanistic understanding and inform future efforts in engineering plants with more efficient Fe uptake or transport systems.

FIT, a regulatory hub for iron deficiency and stress signaling in roots, and FIT-dependent and -independent gene signatures.

Iron (Fe) is vital for plant growth. Plants balance the beneficial and toxic effects of this micronutrient, and tightly control Fe uptake and allocation. Here, we review the role of the basic helix-loop-helix (bHLH) transcription factor FIT (FER-LIKE FE DEFICIENCY-INDUCED TRANSCRIPTION FACTOR) in Fe acquisition. FIT is not only essential, it is also a central regulatory hub in root cells to steer and adjust the rate of Fe uptake by the root in a changing environment. FIT regulates a subset of root Fe deficiency (-Fe) response genes. Based on a combination of co-expression network and FIT-dependent transcriptome analyses, we defined a set of FIT-dependent and FIT-independent gene expression signatures and co-expression clusters that encode specific functions in Fe regulation and Fe homeostasis. These gene signatures serve as markers to integrate novel regulatory factors and signals into the -Fe response cascade. FIT forms a complex with bHLH subgroup Ib transcription factors. Furthermore, it interacts with key regulators from different signaling pathways that either activate or inhibit FIT function to adjust Fe acquisition to growth and environmental constraints. Co-expression clusters and FIT protein interactions suggest a connection of -Fe with ABA responses and root cell elongation processes that can be explored in future studies.

Perioperative course and quality of life in a prospective randomized multicenter phase III trial, comparing standard lobectomy versus anatomical segmentectomy in patients with non-small cell lung cancer up to 2 cm, stage IA (7th edition of TNM staging system).

OBJECTIVES: For early stage non-small cell lung cancer (NSCLC) retrospective data of functionally compromised patients undergoing segmentectomy showed equal outcomes for perioperative complications and quality of life (QoL) compared with lobectomy patients. However no prospectively randomized data comparing patients eligible for both procedures are available. MATERIALS AND METHODS: We conducted a prospective, randomized, multicenter phase III trial and investigated perioperative complications and QoL in patients with NSCLC stage IA (7th edition) undergoing segmentectomy versus lobectomy. The EORTC Questionnaire Core-30 (QLQ C-30) supplemented by thirteen-item lung cancer-specific module (LC13) was assessed before surgery, at discharge, 6 weeks, 3, 6 and 12 months post-surgery. RESULTS: 108 patients with verified or suspected NSCLC up to 2 cm diameter were enrolled, whereby 54 were assigned to lobectomy and 54 to segmentectomy. Due to nodal disease, tumor size and surgical reasons estimated during the operation, eight patients of the segmentectomy group received a lobectomy. In hospital and 90 days mortality was 0% in both groups. Perioperative complications were observed in 6 (11.3%) patients after segmentectomy and in 8 patients (14.8%) after lobectomy (p = 0.563), while the 90-day morbidity were 17% and 25.9% (9 and 14 patients), respectively (p = 0.452). Twelve months after surgery, there was a significant deterioration to the baselines of physical (p < 0.001) and cognitive functioning (p = 0.025), dyspnea (p < 0.001) and fatigue (p = 0.003) in the lobectomy group. Dyspnea showed a faster recovery in the segmentectomy compared to lobectomy group with statistical significance (p = 0.016 after 12 months). CONCLUSION: In patients with early-stage NSCLC, segmentectomy is associated with a statistically not significant lower perioperative morbidity and appears to provide a superior recovery in QoL compared with lobectomy patients.