Integrated Seed Proteome and Phosphoproteome Analyses Reveal Interplay of Nutrient Dynamics,Carbon–Nitrogen Partitioning,and Oxidative Signaling in Chickpea

Nutrient dynamics in storage organs is a complex developmental process that requires coordinated interactions of environmental, biochemical, and genetic factors. Although sink organ developmental events have been identified, understanding of translational and post‐translational regulation of reserve synthesis, accumulation, and utilization in legumes is limited. To understand nutrient dynamics during embryonic and cotyledonary photoheterotrophic transition to mature and germinating autotrophic seeds, an integrated proteomics and phosphoproteomics study in six sequential seed developmental stages in chickpea is performed. MS/MS analyses identify 109 unique nutrient‐associated proteins (NAPs) involved in metabolism, storage and biogenesis, and protein turnover. Differences and similarities in 60 nutrient‐associated phosphoproteins (NAPPs) containing 93 phosphosites are compared with NAPs. Data reveal accumulation of carbon–nitrogen metabolic and photosynthetic proteoforms during seed filling. Furthermore, enrichment of storage proteoforms and protease inhibitors is associated with cell expansion and seed maturation. Finally, combined proteoforms network analysis identifies three significant modules, centered around malate dehydrogenase, HSP70, triose phosphate isomerase, and vicilin. Novel clues suggest that ubiquitin–proteasome pathway regulates nutrient reallocation. Second, increased abundance of NAPs/NAPPs related to oxidative and serine/threonine signaling indicates direct interface between redox sensing and signaling during seed development. Taken together, nutrient signals act as metabolic and differentiation determinant governing storage organ reprogramming.     

不同内固定方式治疗老年踝关节骨折的临床研究

摘要 目的：探讨老年踝关节骨折患者的内固定方式的选择情况及不同内固定方式的疗效,进而指导临床医师根据患者的具体情况选择合适的内固定方式。方法：本研究为回顾性研究,选取我院2016年1月~2018年12月期间收治的老年踝关节骨折患者40例作为研究对象,统计患者一般情况,内容包括骨折块情况、骨折类型、骨质疏松情况、软组织情况与体质。术后随访12个月,评价所有患者末次随访时的踝关节跖屈度、踝关节背伸度、美国足踝外科协会（AOFAS）踝-后足功能评分,记录所有患者的骨折愈合时间。结果：40例研究对象中,使用克氏针张力带11例,Herbert螺钉10例,解剖锁定钢板7例,解剖复合钢板6例,1/3管型钢板6例。骨折块较小、外踝撕脱性骨折的患者主要应用克氏针张力带;伴有骨质疏松的患者主要应用解剖锁定钢板;软组织条件不佳或受损的患者主要应用Herbert螺钉或1/3管型钢板;超重或肥胖患者主要应用解剖复合钢板;瘦弱患者主要应用1/3管型钢板。末次随访时,5种内固定方式患者的踝关节背伸度、踝关节跖屈度、AOFAS踝-后足功能评分比较未见显著性差异（P＞0.05）。5种内固定方式的骨折愈合时间对比差异存在统计学意义（P＜0.05）。结论：老年踝关节骨折应根据患者具体情况选择合理的内固定方式,不同内固定方式患者的骨折愈合时间虽存在差异,但最终均可获得较为满意的疗效。     

不同经皮克氏针内固定术治疗儿童GartlandⅡ、Ⅲ型肱骨髁上骨折疗效对比的回顾性分析

摘要 目的：对比不同经皮克氏针内固定术治疗GartlandⅡ、Ⅲ型肱骨髁上骨折患儿的治疗效果。方法：回顾性分析2017年3月~2020年3月期间我院收治的90例GartlandⅡ、Ⅲ型肱骨髁上骨折患儿的临床资料,根据治疗方式的不同,将患儿分为A组47例（采用切开复位经皮克氏针内固定术治疗）和B组43例（采用闭合复位经皮克氏针内固定术治疗）。观察两组疗效情况、围术期指标情况及并发症发生率。结果：A组、B组的优良率组间对比无统计学差异（P>0.05）。B组的手术时间、骨折愈合时间、住院时间均短于A组,术中失血量少于A组,组间对比差异有统计学意义（P<0.05）。两组术后并发症发生率比较差异无统计学意义（P>0.05）。结论：GartlandⅡ、Ⅲ型肱骨髁上骨折患儿采用闭合复位或切开复位经皮克氏针内固定术均可获得良好的手术效果,其中闭合复位在缩短手术时间、骨折愈合时间、住院时间和减少术中失血量方面更具备优势。     

The effects of hydrocarbon pollution from a two‐stroke outboard engine on the feeding behaviour of Lythrypnus dalli (Perciformes: Gobidae)

Feeding behaviour (time of attack on flake fish food) in Lythrypnus dalli was measured under pollution concentrations of 2.1 and 4.01mgl^?1 total extractable material produced by a two‐stroke outboard engine. The variance of the times increased in pollution versus control treatments for both concentrations. No significant difference was found between the mean times of the control and 2.1 mgl^?1 pollution treatments. A significant difference was found between the mean times of the control and pollution treatments for two different tests at 4.01 mgl^?1. The combination of the increase in mean time of food attack and the increase of variance indicates that pollution concentrations lower than LD50 levels change feeding behaviour, and thus may indirectly effect the survival of the organism.     

Studies on the regioselectivity and kinetics of the action of trypsin on proinsulin and its derivatives using mass spectrometry

Human M-proinsulin was cleaved by trypsin at the R³¹R³²–E³³ and K⁶⁴R⁶⁵–G⁶⁶ bonds (B/C and C/A junctions), showing the same cleavage specificity as exhibited by prohormone convertases 1 and 2 respectively. Buffalo/bovine M-proinsulin was also cleaved by trypsin at the K⁵⁹R⁶⁰–G⁶¹ bond but at the B/C junction cleavage occurred at the R³¹R³²–E³³ as well as the R³¹–R³²E³³ bond. Thus, the human isoform in the native state, with a 31 residue connecting C-peptide, seems to have a unique structure around the B/C and C/A junctions and cleavage at these sites is predominantly governed by the structure of the proinsulin itself. In the case of both the proinsulin species the cleavage at the B/C junction was preferred (65%) over that at the C/A junction (35%) supporting the earlier suggestion of the presence of some form of secondary structure at the C/A junction. Proinsulin and its derivatives, as natural substrates for trypsin, were used and mass spectrometric analysis showed that the k_cat./K_m values for the cleavage were most favourable for the scission of the bonds at the two junctions (1.02 ± 0.08 × 10⁵ s^− 1 M^− 1) and the cleavage of the K²⁹–T³⁰ bond of M-insulin-RR (1.3 ± 0.07 × 10⁵ s^− 1 M^− 1). However, the K²⁹–T³⁰ bond in M-insulin, insulin as well as M-proinsulin was shielded from attack by trypsin (k_cat./K_m values around 1000 s^− 1 M^− 1). Hence, as the biosynthetic path follows the sequence; proinsulin → insulin-RR → insulin, the K²⁹–T³⁰ bond becomes shielded, exposed then shielded again respectively.     

Putative osmosensor – OsHK3b – a histidine kinase protein from rice shows high structural conservation with its ortholog AtHK1 from Arabidopsis

Prokaryotes and eukaryotes respond to various environmental stimuli using the two-component system (TCS). Essentially, it consists of membrane-bound histidine kinase (HK) which senses the stimuli and further transfers the signal to the response regulator, which in turn, regulates expression of various target genes. Recently, sequence-based genome wide analysis has been carried out in Arabidopsis and rice to identify all the putative members of TCS family. One of the members of this family i.e. AtHK1, (a putative osmosensor, hybrid-type sensory histidine kinase) is known to interact with AtHPt1 (phosphotransfer proteins) in Arabidopsis. Based on predicted rice interactome network (PRIN), the ortholog of AtHK1 in rice, OsHK3b, was found to be interacting with OsHPt2. The analysis of amino acid sequence of AtHK1 showed the presence of transmitter domain (TD) and receiver domain (RD), while OsHK3b showed presence of three conserved domains namely CHASE (signaling domain), TD, and RD. In order to elaborate on structural details of functional domains of hybrid-type HK and phosphotransfer proteins in both these genera, we have modeled them using homology modeling approach. The structural motifs present in various functional domains of the orthologous proteins were found to be highly conserved. Binding analysis of the RD domain of these sensory proteins in Arabidopsis and rice revealed the role of various residues such as histidine in HPt protein which are essential for their interaction.     

Cytokinin‐dependent specification of the functional megaspore in the Arabidopsis female gametophyte

The life cycle of higher plants alternates between the diploid sporophytic and the haploid gametophytic phases. In angiosperms, male and female gametophytes develop within the sporophyte. During female gametophyte (FG) development, a single archesporial cell enlarges and differentiates into a megaspore mother cell, which then undergoes meiosis to give rise to four megaspores. In most species of higher plants, including Arabidopsis thaliana, the megaspore closest to the chalaza develops into the functional megaspore (FM), and the remaining three megaspores degenerate. Here, we examined the role of cytokinin signaling in FG development. We characterized the FG phenotype in three triple mutants harboring non‐overlapping T–DNA insertions in cytokinin AHK receptors. We demonstrate that even the strongest mutant is not a complete null for the cytokinin receptors. Only the strongest mutant displayed a near fully penetrant disruption of FG development, and the weakest triple ahk mutant had only a modest FG phenotype. This suggests that cytokinin signaling is essential for FG development, but that only a low threshold of signaling activity is required for this function. Furthermore, we demonstrate that there is elevated cytokinin signaling localized in the chalaza of the ovule, which is enhanced by the asymmetric localization of cytokinin biosynthetic machinery and receptors. We show that an FM‐specific marker is absent in the multiple ahk ovules, suggesting that disruption of cytokinin signaling elements in Arabidopsis blocks the FM specification. Together, this study reveals a chalazal‐localized sporophytic cytokinin signal that plays an important role in FM specification in FG development.