Stent Implantation for Effective Treatment of Refractory Chylothorax due to Superior Vena Cava Obstruction as a Complication of Congenital Cardiac Surgery

Chylothorax is a serious complication of congenital cardiac surgery and is significantly associated with increased morbidity and mortality. Central venous obstruction, which is often related to the insertion of central venous catheters for postoperative management, is known to be an important risk factor for treatment failure and mortality associated with this condition. We present the case of a 6-month-old girl with refractory chylothorax after surgical repair of tetralogy of Fallot. The chylous drainage continued for more than 2 months despite maximal conservative therapy (water restriction, total parenteral nutrition, and infusion of somatostatin and steroid) and surgical ligation of the thoracic duct. Subsequently, we observed stenosis of the superior vena cava (SVC) caused by large thrombi possibly associated with the prolonged use of central venous catheter placed in the internal jugular vein. Because transcatheter balloon dilation failed to relieve the stenosis, we performed stent implantation for the SVC and innominate vein. After the procedure, chylous drainage dramatically reduced, and the patient was discharged from the hospital. In conclusion, central venous obstruction due to thrombosis should be routinely examined when chylothorax is diagnosed and is resistant to conservative therapy after congenital heart surgery. Stent implantation can effectively relieve the venous obstruction and thus be a life-saving treatment option for this difficult condition

Critical Point Mutations for Hepatitis C Virus NS3 Proteinase

AbstractThe hepatitis C virus NS3 proteinase plays an essential role in processing of HCV nonstructural precursor polyprotein. To detect its processing activity, we developed a simpletrans-cleavage assay. Two recombinant plasmids expressing the NS3 proteinase region and a chimeric substrate polyprotein containing the NS5A/5B cleavage site between maltose binding protein and protein A were co-introduced intoEscherichia colicells. The proteinase processed the substrate at the single site during their polyprotein expression. Deletion analysis indicated that the functionally minimal domain of the NS3 proteinase was composed of 146 amino acids, 1059 to 1204. We isolated several cDNA clones encoding the functional domain of the NS3 proteinase from the sera of patients chronically infected with HCV and determined their proteinase activity by thistrans-cleavage assay. Both active and inactive clones existed in the same patients. Comparative sequence analyses of these clones suggested that certain point mutations seemed to be related to the loss of proteolytic activity. This was confirmed by back mutation experiments. Among the critical mutations, Pro-1168 to Thr and Arg-1135 to Gly were intriguing. These amino acids, which are situated near the oxyanion hole, seem to be essential for maintaining the conformation of the active center of the NS3 proteinase

DDA3 recruits microtubule depolymerase Kif2a to spindle poles and controls spindle dynamics and mitotic chromosome movement

Dynamic turnover of the spindle is a driving force for chromosome congression and segregation in mitosis. Through a functional genomic analysis, we identify DDA3 as a previously unknown regulator of spindle dynamics that is essential for mitotic progression. DDA3 depletion results in a high frequency of unaligned chromosomes, a substantial reduction in tension across sister kinetochores at metaphase, and a decrease in the velocity of chromosome segregation at anaphase. DDA3 associates with the mitotic spindle and controls microtubule (MT) dynamics. Mechanistically, DDA3 interacts with the MT depolymerase Kif2a in an MT-dependent manner and recruits Kif2a to the mitotic spindle and spindle poles. Depletion of DDA3 increases the steady-state levels of spindle MTs by reducing the turnover rate of the mitotic spindle and by increasing the rate of MT polymerization, which phenocopies the effects of partial knockdown of Kif2a. Thus, DDA3 represents a new class of MT-destabilizing protein that controls spindle dynamics and mitotic progression by regulating MT depolymerases

Plk1- and β-TrCP–dependent degradation of Bora controls mitotic progression

Through a convergence of functional genomic and proteomic studies, we identify Bora as a previously unknown cell cycle protein that interacts with the Plk1 kinase and the SCF–β-TrCP ubiquitin ligase. We show that the Bora protein peaks in G2 and is degraded by proteasomes in mitosis. Proteolysis of Bora requires the Plk1 kinase activity and is mediated by SCF–β-TrCP. Plk1 phosphorylates a conserved DSGxxT degron in Bora and promotes its interaction with β-TrCP. Mutations in this degron stabilize Bora. Expression of a nondegradable Bora variant prolongs the metaphase and delays anaphase onset, indicating a physiological requirement of Bora degradation. Interestingly, the activity of Bora is also required for normal mitotic progression, as knockdown of Bora activates the spindle checkpoint and delays sister chromatid segregation. Mechanistically, Bora regulates spindle stability and microtubule polymerization and promotes tension across sister kinetochores during mitosis. We conclude that tight regulation of the Bora protein by its synthesis and degradation is critical for cell cycle progression

An Arabidopsis SBP-domain fragment with a disrupted C-terminal zinc-binding site retains its tertiary structure

AbstractSQUAMOSA promoter-binding proteins (SBPs) form a major family of plant-specific transcription factors, mainly related to flower development. SBPs share a highly conserved DNA-binding domain of ∼80 amino acids (SBP domain), which contains two non-interleaved zinc-binding sites formed by eight conserved Cys or His residues. In the present study, an Arabidopsis SPL12 SBP-domain fragment that lacks a Cys residue involved in the C-terminal zinc-binding pocket was found to retain a folded structure, even though only a single Zn2+ ion binds to the fragment. Solution structure of this fragment determined by NMR is very similar to the previously determined structures of the full SBP domains of Arabidopsis SPL4 and SPL7. Considering the previous observations that chelating all the Zn2+ ions of SBPs resulted in the complete unfolding of the structure and that a mutation of the Cys residue equivalent to that described above impaired the DNA-binding activity, we propose that the Zn2+ ion at the N-terminal site is necessary to maintain the overall tertiary structure, while the Zn2+ ion at the C-terminal site is necessary for the DNA binding, mainly by guiding the basic C-terminal loop to correctly fit into the DNA groove

Effect of Charge Substitutions at Residue His-142 on Voltage Gating of Connexin43 Channels

AbstractPrevious studies indicate that the carboxyl terminal of connexin43 (Cx43CT) is involved in fast transjunctional voltage gating. Separate studies support the notion of an intramolecular association between Cx43CT and a region of the cytoplasmic loop (amino acids 119–144; referred to as “L2”). Structural analysis of L2 shows two α-helical domains, each with a histidine residue in its sequence (H126 and H142). Here, we determined the effect of H142 replacement by lysine, alanine, and glutamate on the voltage gating of Cx43 channels. Mutation H142E led to a significant reduction in the frequency of occurrence of the residual state and a prolongation of dwell open time. Macroscopically, there was a large reduction in the fast component of voltage gating. These results resembled those observed for a mutant lacking the carboxyl terminal (CT) domain. NMR experiments showed that mutation H142E significantly decreased the Cx43CT-L2 interaction and disrupted the secondary structure of L2. Overall, our data support the hypothesis that fast voltage gating involves an intramolecular particle-receptor interaction between CT and L2. Some of the structural constrains of fast voltage gating may be shared with those involved in the chemical gating of Cx43