Phosphorylation screening identifies translational initiation factor 4GII as an intracellular target of Ca(2+)/calmodulin-dependent protein kinase I

CaMKI is a Ca2+/calmodulin-dependent protein kinase that is widely expressed in eukaryotic cells and tissues but for which few, if any, physiological substrates are known. We screened a human lung cDNA expression library for potential CaMKI substrates by solid phase in situ phosphorylation ("phosphorylation screening"). Multiple overlapping partial length cDNAs encoding three proteins were detected. Two of these proteins are known: 6-phosphofructo-2-kinase/fructose 2,6-bisphosphatase and eukaryotic translation initiation factor (eIF) 4GII. To determine whether CaMKI substrates identified by phosphorylation screening represent authentic physiological targets, we examined the potential for [Ca2+]i- and CaMKI-dependent phosphorylation of eIF4GII in vitro and in vivo. Endogenous eIF4GII immunoprecipitated from HEK293T cells was phosphorylated by CaMKI, in vitro as was a recombinant fragment of eIF4GII encompassing the central and C-terminal regions. The latter phosphorylation occurred with favorable kinetics (Km = 1 microm; kcat = 1.8 s-1) at a single site, Ser1156, located in a segment of eIF4GII aligning with the phosphoregion of eIF4GI. Phosphopeptide mapping and back phosphorylation experiments revealed [Ca2+]i-dependent, CaMKI site-specific, eIF4GII phosphorylation in vivo. This phosphorylation was blocked by kinase-negative CaMKI consistent with a requirement for endogenous CaMKI for in vivo eIF4GII phosphorylation. We conclude that phosphorylation screening is an effective method for searching for intracellular targets of CaMKI and may have identified a new role of Ca2+ signaling to the translation apparatus.     

Metabolic engineering of Corynebacterium glutamicum for increasing the production of l-ornithine by increasing NADPH availability

The experiments presented here were based on the conclusions of our previous proteomic analysis. Increasing the availability of glutamate by overexpression of the genes encoding enzymes in the l-ornithine biosynthesis pathway upstream of glutamate and disruption of speE, which encodes spermidine synthase, improved l-ornithine production by Corynebacterium glutamicum. Production of l-ornithine requires 2 moles of NADPH per mole of l-ornithine. Thus, the effect of NADPH availability on l-ornithine production was also investigated. Expression of Clostridium acetobutylicum gapC, which encodes NADP-dependent glyceraldehyde-3-phosphate dehydrogenase, and Bacillus subtilis rocG, which encodes NAD-dependent glutamate dehydrogenase, led to an increase of l-ornithine concentration caused by greater availability of NADPH. Quantitative real-time PCR analysis demonstrates that the increased levels of NADPH resulted from the expression of the gapC or rocG gene rather than that of genes (gnd, icd, and ppnK) involved in NADPH biosynthesis. The resulting strain, C. glutamicum ΔAPRE::rocG, produced 14.84 g l^?1 of l-ornithine. This strategy of overexpression of gapC and rocG will be useful for improving production of target compounds using NADPH as reducing equivalent within their synthetic pathways.     

High-quality genome assembly of Huazhan and Tianfeng,the parents of an elite rice hybrid Tian-you-hua-zhan

High-quality rice reference genomes have accelerated the comprehensive identification of genome-wide variations and research on functional genomics and breeding. Tian-you-hua-zhan has been a leading hybrid in China over the past decade. Here, de novo genome assembly strategy optimization for the rice indica lines Huazhan (HZ) and Tianfeng (TF), including sequencing platforms, assembly pipelines and sequence depth, was carried out. The PacBio and Nanopore platforms for long-read sequencing were utilized, with the Canu, wtdbg2, SMARTdenovo, Flye, Canu-wtdbg2, Canu-SMARTdenovo and Canu-Flye assemblers. The combination of PacBio and Canu was optimal, considering the contig N50 length, contig number, assembled genome size and polishing process. The assembled contigs were scaffolded with Hi-C data, resulting in two “golden quality” rice reference genomes, and evaluated using the scaffold N50, BUSCO, and LTR assembly index. Furthermore, 42,625 and 41,815 non-transposable element genes were annotated for HZ and TF, respectively. Based on our assembly of HZ and TF, as well as Zhenshan97, Minghui63, Shuhui498 and 9311, comprehensive variations were identified using Nipponbare as a reference. The de novo assembly strategy for rice we optimized and the “golden quality” rice genomes we produced for HZ and TF will benefit rice genomics and breeding research, especially with respect to uncovering the genomic basis of the elite traits of HZ and TF.

     

Nosocomial Infection in Adult Admissions with Hematological Malignancies Originating from Different Lineages: A Prospective Observational Study

Background

Nosocomial infection (NI) causes prolonged hospital stays, increased healthcare costs, and higher mortality among patients with hematological malignancies (HM). However, few studies have compared the incidence of NI according to the HM lineage.

Objective

To compare the incidence of NI according to the type of HM lineage, and identify the risk factors for NI.

Methods

This prospective observational study monitored adult patients with HM admitted for >48 hours to the General Hospital of the People''s Liberation Army during 2010–2013. Attack rates and incidences of NI were compared, and multivariable logistic regression was used to control for confounding effects.

Results

This study included 6,613 admissions from 1,922 patients. During these admissions, 1,023 acquired 1,136 NI episodes, with an attack rate of 15.47% and incidence of 9.6‰ (95% CI: 9.1–10.2). Higher rates and densities of NIs were observed among myeloid neoplasm (MN) admissions, compared to lymphoid neoplasm (LN) admissions (28.42% vs. 11.00%, P<0.001 and 11.4% vs. 8.4‰, P<0.001). NI attack rates in acute myeloid leukemia (AML) and myelodysplastic/myeloproliferative neoplasm (MDS/MPN) were higher than those in MDS (30.69% vs. 20.19%, P<0.001; 38.89% vs. 20.19%, P = 0.003). Attack rates in T/NK-cell neoplasm and B-cell neoplasm were higher than those in Hodgkin lymphoma (15.04% vs. 3.65%; 10.94% vs. 3.65%, P<0.001). Multivariable regression analysis indicated prolonged hospitalization, presence of central venous catheterization, neutropenia, current stem cell transplant, infection on admission, and old age were independently associated with higher NI incidence. After adjusting for these factors, MN admissions still had a higher risk of infection (odds ratio 1.34, 95% CI: 1.13–1.59, P<0.001).

Conclusion

Different NI attack rates were observed for HM from different lineages, with MN lineages having a higher attack rate and incidence than LN lineages. Special attention should be paid to MN admissions, especially AML and MDS/MPN admissions, to control NI incidence.     

超低频脉冲磁场抑制癌瘤和提高细胞免疫功能的实验研究

在报道用电子显微镜观测超低频脉冲磁场(峰值磁场0.6～2.0T,磁场梯度10～100T·m^-1,脉冲宽度20～200ms,重复频率0.16～1.34Hz)抑制鼠S-180肉瘤和加强免疫细胞溶癌作用以后,报道了用Faulgen染色法测定肉瘤细胞核的DNA倍性,和用电镜技术和细胞结构的体视学分析磁场对癌细胞形态的影响,观测到磁场影响癌细胞的代谢:磁场使癌细胞的恶性程度降低,抑制其高速和异形生长;磁场抑制癌细胞的分裂和DNA的复制;磁场提高细胞免疫功能,加强淋巴细胞、浆细胞反应.     

Diminished AMPK signaling response to fasting in thioredoxin-interacting protein knockout mice

Thioredoxin-interacting protein (Txnip) knockout (TKO) mice exhibit impaired response to fasting. Herein, we showed that activation of adenine monophosphate-activated protein kinase and cellular AMP levels were diminished in the heart and soleus muscle but not in gastrocnemius muscle of fasting TKO mice. Similarly, glycogen content in fasted TKO mice was increased in oxidative muscles but was not different in glycolytic muscles. These data suggest Txnip deficiency has a higher impact on oxidative muscle than glycolytic muscles and provide new insights into the metabolic role of Txnip.     

老年腰椎骨折术后深部切口感染患者病原菌和感染危险因素分析

分析老年腰椎骨折术后切口深部感染患者的病原菌分布特征及感染危险因素,为该类患者的治疗提供参考。

选择2018年2月至2021年2月于我院行腰椎手术治疗且符合纳入标准的患者589例,记录患者感染发生率,并对感染病原菌分布特征及感染的危险因素进行单因素和多因素分析,同时探讨相应对策。

589例腰椎骨折手术患者有41例（6.96%）出现切口深部感染。41例感染患者的渗出液或分泌物中鉴定分离出53株病原菌,其中30例患者为单菌株感染,占73.17%;11例患者为多菌株感染,占26.83%。53株病原菌中革兰阴性菌30株,革兰阳性菌22株,真菌1株。革兰阴性菌中,大肠埃希菌对哌拉西林的耐药率最高,为91.67%;对美罗培南、亚胺培南及厄他培南的耐药率相对较低,为16.67%;铜绿假单胞菌对头孢吡肟、头孢曲松、头孢唑林和复方磺胺甲噁唑的耐药率最高,为90.00%;对环丙沙星和阿米卡星的耐药率相对较低,为20.00%。革兰阳性菌中金黄色葡萄球菌对青霉素和红霉素的耐药率最高,为76.92%;对万古霉素、利奈唑胺的耐药率为0.00%。术后感染患者与非感染患者在年龄分布、BMI、是否合并糖尿病、手术持续时间、是否输血等方面的差异有统计学意义（均P＜0.05）;在性别、是否存在烟酒史、是否合并高血压或冠心病、术中失血量等方面差异无统计学意义（均P＞0.05）。Logistic回归分析显示,年龄≥60岁、BMI≥28 kg/m²、合并糖尿病、手术时间≥2 h及预防性应用抗生素为腰椎骨折术后患者切口深部感染的独立危险因素（均P＜0.05）。

腰椎骨折术后患者深部切口感染病原菌种类较多,以金黄色葡萄球菌和大肠埃希菌为主,且药敏性普遍较低。年龄≥60岁、BMI≥28 kg/m²、合并糖尿病、手术时间≥2 h及预防性应用抗生素为腰椎骨折术后患者切口深部感染的独立危险因素,临床应根据药敏情况合理选择抗菌药物以降低感染发生率。

     

Epitope Tags beside the N-Terminal Cytoplasmic Tail of Human BST-2 Alter Its Intracellular Trafficking and HIV-1 Restriction

BST-2 blocks the particle release of various enveloped viruses including HIV-1, and this antiviral activity is dependent on the topological arrangement of its four structural domains. Several functions of the cytoplasmic tail (CT) of BST-2 have been previously discussed, but the exact role of this domain remains to be clearly defined. In this study, we investigated the impact of truncation and commonly-used tags addition into the CT region of human BST-2 on its intracellular trafficking and signaling as well as its anti-HIV-1 function. The CT-truncated BST-2 exhibited potent inhibition on Vpu-defective HIV-1 and even wild-type HIV-1. However, the N-terminal HA-tagged CT-truncated BST-2 retained little antiviral activity and dramatically differed from its original protein in the cell surface level and intracellular localization. Further, we showed that the replacement of the CT domain with a hydrophobic tag altered BST-2 function possibly by preventing its normal vesicular trafficking. Notably, we demonstrated that a positive charged motif “KRXK” in the conjunctive region between the cytotail and the transmembrane domain which is conserved in primate BST-2 is important for the protein trafficking and the antiviral function. These results suggest that although the CT of BST-2 is not essential for its antiviral activity, the composition of residues in this region may play important roles in its normal trafficking which subsequently affected its function. These observations provide additional implications for the structure-function model of BST-2.