Modulation of Intracellular Calcium Levels by Calcium Lactate Affects Colon Cancer Cell Motility through Calcium-Dependent Calpain

Cancer cell motility is a key phenomenon regulating invasion and metastasis. Focal adhesion kinase (FAK) plays a major role in cellular adhesion and metastasis of various cancers. The relationship between dietary supplementation of calcium and colon cancer has been extensively investigated. However, the effect of calcium (Ca²⁺) supplementation on calpain-FAK-motility is not clearly understood. We sought to identify the mechanism of FAK cleavage through Ca²⁺ bound lactate (CaLa), its downstream signaling and role in the motility of human colon cancer cells. We found that treating HCT116 and HT-29 cells with CaLa immediately increased the intracellular Ca²⁺ (iCa²⁺) levels for a prolonged period of time. Ca²⁺ influx induced cleavage of FAK into an N-terminal FAK (FERM domain) in a dose-dependent manner. Phosphorylated FAK (p-FAK) was also cleaved in to its p-N-terminal FAK. CaLa increased colon cancer cells motility. Calpeptin, a calpain inhibitor, reversed the effects of CaLa on FAK and pFAK cleavage in both cancer cell lines. The cleaved FAK translocates into the nucleus and modulates p53 stability through MDM2-associated ubiquitination. CaLa-induced Ca²⁺ influx increased the motility of colon cancer cells was mediated by calpain activity through FAK and pFAK protein destabilization. In conclusion, these results suggest that careful consideration may be given in deciding dietary Ca²⁺ supplementation to patient undergoing treatment for metastatic cancer.     

Draft Genome Sequence of a Novel Bacterial Strain,LSJC7, Belonging to the Family Enterobacteriaceae with Dual Resistance to Arsenic and Tetracycline

Strain LSJC7, with dual resistance to arsenic and tetracycline, was isolated from an antimony tailing in China. Its 16S rRNA gene sequence has the highest similarity to that of Enterobacter cloacae subsp. dissolvens LMG 2683^T (97.02%). Here we present the approximately 4.6-Mbp draft genome sequence of strain LSJC7.     

The Tumor Suppressor pVHL Down-regulates Never-in-Mitosis A-related Kinase 8 via Hypoxia-inducible Factors to Maintain Cilia in Human Renal Cancer Cells

NEK8 (never in mitosis gene A (NIMA)-related kinase 8) is involved in cytoskeleton, cilia, and DNA damage response/repair. Abnormal expression and/or dysfunction of NEK8 are related to cancer development and progression. However, the mechanisms that regulate NEK8 are not well declared. We demonstrated here that pVHL may be involved in regulating NEK8. We found that CAK-I cells with wild-type vhl expressed a lower level of NEK8 than the cells loss of vhl, such as 786-O, 769-P, and A-498 cells. Moreover, pVHL overexpression down-regulated the NEK8 protein in 786-O cells, whereas pVHL knockdown up-regulated NEK8 in CAK-I cells. In addition, we found that the positive hypoxia response elements (HREs) are located in the promoter of the nek8 sequence and hypoxia could induce nek8 expression in different cell types. Consistent with this, down-regulation of hypoxia-inducible factors α (HIF-1α or HIF-2α) by isoform-specific siRNA reduced the ability of hypoxia inducing nek8 expression. In vivo, NEK8 and HIF-1α expression were increased in kidneys of rats subjected to an experimental hypoxia model of ischemia and reperfusion. Furthermore, NEK8 siRNA transfection significantly blocked pVHL-knockdown-induced cilia disassembling, through impairing the pVHL-knockdown-up-regulated NEK8 expression. These results support that nek8 may be a novel hypoxia-inducible gene. In conclusion, our findings show that nek8 may be a new HIF target gene and pVHL can down-regulate NEK8 via HIFs to maintain the primary cilia structure in human renal cancer cells.     

Association between misalignment of circadian rhythm and obesity in Korean men: Sixth Korea National Health and Nutrition Examination Survey

ABSTRACT

Background: The disruption of circadian rhythm has been found to associate with obesity in vivo and in vitro. Sleep duration, eating habits, total feeding time, and nightshift work can also affect circadian rhythms. This study investigated the association between misalignment of circadian rhythm and obesity in Korean men, using a cross-sectional database.

Methods: This study used data from the Korean National Health and Nutrition Examination Survey (KNHANES), whose study population was 3,658 men aged 18 to 60 years. General and abdominal obesity was defined as a body mass index (BMI) ≥ 25 kg/m² and waist circumference ≥ 90 cm, respectively. Circadian rhythm factors were determined with a self-report questionnaire and included breakfast frequency, sleep duration, and work time. Frequency of breakfast was divided into regular breakfast (five to seven times a week) and irregular breakfast (less than five times a week). Sleep duration was divided into less than 7 hours, 7–9 hours, and over 9 hours. Working time was defined as day/evening, night shift, and other type. The adjusted odds ratios (aORs) and 95% confidence intervals (CIs) for general and abdominal obesity were calculated using multivariable logistic regression according to the number of factors that disturb the circadian rhythm.

Results: Participants with 1 (aOR 1.34, 95% Cl 1.10–1.61) and ≥2 (aOR 1.62, 95% Cl 1.29–2.05) factors disturbing circadian rhythms were associated with elevated risk for general obesity. Similarly, those with 1 (aOR 1.33, 95% Cl 1.09–1.63) and ≥2 (aOR 1.70, 95% Cl 1.32–2.20) factors had elevated risk for abdominal obesity.

Conclusions: Factors disturbing the circadian rhythm were associated with general and abdominal obesity. Additional studies are needed, and associations with metabolic diseases should be investigated.     

UHRF2, a Ubiquitin E3 Ligase,Acts as a Small Ubiquitin-like Modifier E3 Ligase for Zinc Finger Protein 131

Small ubiquitin-like modifier (SUMO), a member of the ubiquitin-related protein family, is covalently conjugated to lysine residues of its substrates in a process referred to as SUMOylation. SUMOylation occurs through a series of enzymatic reactions analogous to that of the ubiquitination pathway, resulting in modification of the biochemical and functional properties of substrates. To date, four mammalian SUMO isoforms, a single heterodimeric SUMO-activating E1 enzyme SAE1/SAE2, a single SUMO-conjugating E2 enzyme ubiquitin-conjugating enzyme E2I (UBC9), and a few subgroups of SUMO E3 ligases have been identified. Several SUMO E3 ligases such as topoisomerase I binding, arginine/serine-rich (TOPORS), TNF receptor-associated factor 7 (TRAF7), and tripartite motif containing 27 (TRIM27) have dual functions as ubiquitin E3 ligases. Here, we demonstrate that the ubiquitin E3 ligase UHRF2 also acts as a SUMO E3 ligase. UHRF2 effectively enhances zinc finger protein 131 (ZNF131) SUMOylation but does not enhance ZNF131 ubiquitination. In addition, the SUMO E3 activity of UHRF2 on ZNF131 depends on the presence of SET and RING finger-associated and nuclear localization signal-containing region domains, whereas the critical ubiquitin E3 activity RING domain is dispensable. Our findings suggest that UHRF2 has independent functional domains and regulatory mechanisms for these two distinct enzymatic activities.     

Pre‐copulation intervals,copulation frequencies,and initial progeny sex ratios in two biotypes of whitefly,Bemisia tabaci

The whitefly Bemisia tabaci (Gennadius) (Homoptera: Aleyrodidae) is a species complex, and its systematic classification requires controlled crossing experiments among its genetic groups. Accurate information on pre‐copulation intervals, copulation frequencies, and initial frequency of egg fertilization of newly emerged adults is critical for designing procedures for collecting the virgin adults necessary for these experiments. In the literature, considerable variation is reported between B. tabaci populations, with respect to the length of the pre‐copulation interval and the initial frequency of egg fertilization. Here, we used a video‐recording method to observe continuously the copulation behaviour of the Mediterranean/Asia Minor/Africa (B biotype) and the Asia II (ZHJ1 biotype) groups of B. tabaci. We also recorded the initial frequency of egg fertilization, as determined by the sex of the progeny. When adults were caged in female–male pairs on leaves of cotton plants, the earliest copulation events occurred 2–6 h after emergence; at 12 h after emergence 56–84% of the females had copulated at least once, and nearly all (92–100%) had copulated at least once by 36 h after emergence. Both females and males copulated repeatedly. Approximately 80 and 20% of copulation events occurred during the photophase and scotophase, respectively. By 72 h post‐emergence, the females of the B and ZHJ1 biotypes had copulated on average 6.1 and 3.9 times, respectively. When adults were caged in groups on plants 1–13 h after emergence, 30–35% of the eggs deposited during this period were fertilized, and approximately 90% of females were fertilized by the end of the 13 h. Although timing of copulation differed in detail between the two genetic groups, the results demonstrate that B. tabaci adults can start to copulate as early as 2–6 h post‐emergence and the majority of females can become fertilized on the day that they emerge.     

Polyphosphate Formation from Pyrophosphate in Intact Cells of a Photosynthetic Bacterium, Rhodospirillum rubrum

The hypothesis that the photosynthetic bacterium, Rhodospirillumrubrum, may synthesize polyphosphate using pyrophosphate asa precursor has been tested. The cells were cultured in thepyrophosphate media, and the cell yield, cell shape, and thepolyphosphate content were determined and compared with thatgrown in the orthophosphate media. In the control experiment, the total amount of polyphosphateformed by the cultures grown in the pyrophosphate media wasfound to be up to 70% less than that grown in orthophosphatemedia. When the ATP formation was blocked by using the phosphorylationinhibitor, antimycin A, however, the cells grown in the pyrophosphatemedia formed up to 50% more polyphosphate than that grown inthe orthophosphate media. The results seem to support the hypothesis that Rhodospirillumrubrum may use pyrophosphate as the precursor of polyphosphate. (Received May 26, 1986; Accepted February 21, 1987)     

金铁锁的一个新三萜成分

金铁锁(Psammosilene tunicoides W.C.Wu et C.Y.Wu)系石竹科金铁锁属单一种植物,产于我国西南部。该植物为传统中药,具止血、止痛、活血等多种功效。其根部富含皂甙成分,有镇痛、抗炎和抑菌等生理活性。我们曾报道了将其根中提取的粗皂甙酸性水解后,分离到的六个齐墩果酸型三萜皂甙元。本文报道从中分离到的另一新三萜成分:3-hydroxy-12,14-diene-27-nordeane-28-nic acid。     

重楼属植物甾体皂甙的高效液相色谱分析

应用高效液相层析技术,对重楼属十八个种植物的甾体皂甙进行了定性、定量分析。植物用甲醇提取,抽出物经DIAION柱,以90％甲醇洗出总甙。在HPLC上,用ODS柱先将总甙以用醇:水(9:1)洗脱分为三馏段,每馏再在ODS柱或Rp-8柱上以甲醇:水(8:2;7:3.5)洗脱,使各个皂甙成分完全分离。被分离的每个色谱峰与已知重楼皂甙的保留时间进行比较并配合HPLC的加入法,TLC分析及用HPLC制备少量样品做MS测定来加以定性鉴定。定量采用内标及校正曲线法。     

Immunocytochemical localization of lactoferrin in human neutrophils

Summary The subcellular localization of lactoferrin in human neutrophils was studied by an electron-microscopic immunoperoxidase method. This molecule was detected in small granules of blood polymorphonuclear leukocytes. A morphometrical analysis showed that there was no significant difference in the mean size between lactoferrin-positive and myeloperoxidase-negative granules. In contrast, the mean size of myeloperoxidase-positive granules was significantly larger than that of lactoferrin-positive granules. This indicates that lactoferrin is contained in the myeloperoxidase-negative, secondary, granules of human neutrophils. In immature bone marrow mononuclear neutrophils, lactoferrin was present in cytoplasmic granules of somewhat larger size than lactoferrin-positive granules of polymorphonuclear leucocytes. A morphometrical study showed that the mean size of lactoferrin-positive granules was significantly greater in immature bone marrow cells than in polymorphonuclear leucocytes. This indicates that lactoferrin-positive granules decrease in size as the cells mature. Besides cytoplasmic granules, lactoferrin was demonstrated in the Golgi complex and a part of the rough endoplasmic reticulum of immature bone marrow neutrophils, probably myelocytes and early metamyelocytes. These results show that lactoferrin is synthesized and packed into secondary granules in immature bone marrow neutrophils and therefore that the secondary granules are a type of secretory granule.