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1.
J Biol Chem ; 295(43): 14780-14788, 2020 10 23.
Article in English | MEDLINE | ID: mdl-32938714

ABSTRACT

The mitotic kinase Aurora B regulates the condensation of chromatin into chromosomes by phosphorylating chromatin proteins during early mitosis, whereas the phosphatase PP1γ performs the opposite function. The roles of Aurora B and PP1γ must be tightly coordinated to maintain chromosomes at a high phosphorylation state, but the precise mechanisms regulating their function remain largely unclear. Here, mainly through immunofluorescence microscopy and co-immunoprecipitation assays, we find that dissociation of PP1γ from chromosomes is essential for maintaining chromosome phosphorylation. We uncover that PP1γ is recruited to mitotic chromosomes by its regulatory subunit Repo-Man in the absence of Aurora B activity and that Aurora B regulates dissociation of PP1γ by phosphorylating and disrupting PP1γ-Repo-Man interactions on chromatin. Overexpression of Repo-Man mutants that cannot be phosphorylated or inhibition of Aurora B kinase activity resulted in the retention of PP1γ on chromatin and prolonged the chromatin condensation process; a similar outcome was caused by the ectopic targeting of PP1γ to chromatin. Together, our findings reveal a novel regulation mechanism of chromatin condensation in which Aurora B counteracts PP1γ activity by releasing PP1γ from Repo-Man and may have important implications for understanding the regulations of dynamic structural changes of the chromosomes in mitosis.


Subject(s)
Aurora Kinase B/metabolism , Carrier Proteins/metabolism , Cell Cycle Proteins/metabolism , Nuclear Proteins/metabolism , Protein Phosphatase 1/metabolism , Chromatin/metabolism , Chromosomes, Human/metabolism , HeLa Cells , Humans , Mitosis , Phosphorylation , Protein Interaction Maps
2.
J Sci Food Agric ; 97(6): 1861-1867, 2017 Apr.
Article in English | MEDLINE | ID: mdl-27507439

ABSTRACT

BACKGROUND: To clarify the formation mechanism of trans linoleic acid isomers in edible oils during the heating process, trilinolein and trilinoelaidin, as representative oils, were placed in glass ampoules and sealed before heating at 180, 240 and 320 °C. The glass ampoules were removed at regular time intervals, and the contents were analyzed by infrared spectroscopy. The samples were then subjected to derivatization into their methyl esters for gas chromatographic analysis. RESULTS: Analysis results show that 9c,12c and 9t,12t fatty acids from trilinolein and trilinoelaidin molecules undergo chemical bond rotation, migration and degradation, leading to the formation of non-conjugated linoleic acids (NLAs), conjugated linoleic acids (CLAs) and aldehydes. The formation rate of isomers from the 9c,12c fatty acid is higher than that of the 9t,12t fatty acid. The production of aldehydes increases with heating temperature and time. The isomerization pathways involved in the formation of NLAs and CLAs during heating are clearly presented. CONCLUSION: These findings suggest possible pathways of NFA and CFA formation from heated trilinolein and trilinoelaidin, complement the mechanistic studies previously published in the literature, and provide a theoretical basis for future control of the quality and safety of fats and oils. © 2016 Society of Chemical Industry.


Subject(s)
Linoleic Acid/chemistry , Plant Oils/chemistry , Triglycerides/chemistry , Chromatography, Gas , Cooking , Hot Temperature , Isomerism , Linoleic Acids, Conjugated/chemistry , Molecular Structure
3.
J Biol Chem ; 290(28): 17546-58, 2015 Jul 10.
Article in English | MEDLINE | ID: mdl-25987563

ABSTRACT

Aurora kinase A and B share great similarity in sequences, structures, and phosphorylation motif, yet they show different localizations and play distinct crucial roles. The factors that determine such differences are largely unknown. Here we targeted Aurora A to the localization of Aurora B and found that Aurora A phosphorylates the substrate of Aurora B and substitutes its function in spindle checkpoint. In return, the centrosome targeting of Aurora B substitutes the function of Aurora A in the mitotic entry. Expressing the chimera proteins of the Auroras with exchanged N termini in cells indicates that the divergent N termini are also important for their spatiotemporal localizations and functions. Collectively, we demonstrate that functional divergence of Aurora kinases is determined by spatial compartmentalization, and their divergent N termini also contribute to their spatial and functional differentiation.


Subject(s)
Aurora Kinase A/metabolism , Aurora Kinase B/metabolism , Amino Acid Sequence , Animals , Aurora Kinase A/chemistry , Aurora Kinase A/genetics , Aurora Kinase B/chemistry , Aurora Kinase B/genetics , Cell Compartmentation , Cell Cycle Checkpoints , Centrosome/metabolism , Chromatin/metabolism , Evolution, Molecular , HeLa Cells , Histones/metabolism , Humans , Kinetochores/metabolism , Mitosis , Models, Biological , Molecular Sequence Data , Phosphorylation , Recombinant Fusion Proteins/chemistry , Recombinant Fusion Proteins/genetics , Recombinant Fusion Proteins/metabolism , Sequence Homology, Amino Acid , Spindle Apparatus/metabolism
4.
Proc Natl Acad Sci U S A ; 110(38): 15295-300, 2013 Sep 17.
Article in English | MEDLINE | ID: mdl-24003142

ABSTRACT

Kinetochore capture by dynamic kinetochore microtubule fibers (K fibers) is essential for proper chromosome alignment and accurate distribution of the replicated genome during cell division. Although this capture process has been extensively studied, the mechanisms underlying the initiation of this process and the proper formation of the K fibers remain largely unknown. Here we show that transforming acidic coiled-coil-containing protein 3 (TACC3) is essential for kinetochore capture and proper K-fiber formation in HeLa cells. To observe the assembly of acentrosomal microtubules more clearly, the cells were released from higher concentrations of nocodazole into zero or lower concentrations. We find that small acentrosomal TACC3-microtubule aster formation near the kinetochores and binding of the asters with the kinetochores are the initial steps of the kinetochore capture by the acentrosomal microtubules, and that the sorting of kinetochore-captured acentrosomal microtubules with centrosomal microtubules leads to the capture of kinetochore by centrosomal microtubules from both spindle poles. We demonstrate that the sorting of the TACC3-associated microtubules with the centrosomal microtubules is a crucial process for spindle assembly and chromosome movement. These findings, which are also supported in the unperturbed mitosis without nocodazole, reveal a critical TACC3-dependent acentrosomal microtubule nucleation and sorting process to regulate kinetochore-microtubule connections and provide deep insight into the mechanisms of mitotic spindle assembly and chromosome alignment.


Subject(s)
Kinetochores/metabolism , Microtubule-Associated Proteins/metabolism , Microtubules/metabolism , Spindle Apparatus/physiology , Blotting, Western , Fluorescent Antibody Technique , HeLa Cells , Humans , RNA Interference , RNA, Small Interfering/genetics , Spindle Apparatus/metabolism
5.
J Food Sci Technol ; 53(3): 1487-95, 2016 Mar.
Article in English | MEDLINE | ID: mdl-27570273

ABSTRACT

The effects of technical cashew nut shell liquid (TCNSL) on the trans isomerization of edible oils during heating are investigated. Edible oils were subjected to thermal treatment at various heating times and temperatures. Our results show that the addition of TCNSL to edible oils at the appropriate concentration during heating suppresses trans fatty acid formation and induces formation of conjugated linoleic acid (CLA) isomers. A concentration of 0.2 % TCNSL demonstrates the best ability to inhibit formation of trans-oleic acid, trans-linoleic acid, and trans-linolenic acid isomers as well as increase the formation of 9 t,11 t-CLA and 10 t,12 t-CLA isomers. Our analysis indicates that the presence of 0.2 % TCNSL in corn oil does not significantly reduce the acid value, but may significantly lower the peroxide value. TCNSL is also observed to have better function compared to Vitamin E (VE) and tertiary butylhydroquinone (TBHQ), indicating that it may be considered an effective additive in edible oils.

6.
Food Chem ; 174: 299-305, 2015 May 01.
Article in English | MEDLINE | ID: mdl-25529684

ABSTRACT

Trilinolein, with or without additives, was placed in glass ampoules and subjected to thermal treatment at 180 °C or 240 °C for 8h. Thermal treatment of trilinolein at 180 °C and 240 °C produced twice the amount of trans nonconjugated linoleic acids (NLAs) compared to conjugated linoleic acids (CLAs), and nitrogen stream reduced the amount of both trans NLA and CLA products. The presence of additives resulted in the suppression or induction of trans NLAs and CLAs, depending on the type of additive, the concentration of the additive, and the heating temperature. Our analysis indicates that TBHQ is an effective additive for reducing trans NLA formation and inducing trans CLA formation in frying oil. Glutathione and L-cysteine at 0.1% may also be used as additives for frying oil. At suitable concentrations, Fe(3+) and Al(3+) ions derived from oils can reduce trans NLAs and induce trans CLAs during frying.


Subject(s)
Trans Fatty Acids/chemistry , Triglycerides/chemistry , Butylated Hydroxyanisole/pharmacology , Butylated Hydroxytoluene/pharmacology , Chromatography, Gas , Cooking , Cysteine/pharmacology , Hot Temperature , Hydroquinones/pharmacology , Isomerism , Linoleic Acid/chemistry , Linoleic Acids, Conjugated/chemistry , Vitamin E/pharmacology
7.
J Cell Biol ; 210(3): 373-83, 2015 Aug 03.
Article in English | MEDLINE | ID: mdl-26240182

ABSTRACT

A steady-state metaphase spindle maintains constant length, although the microtubules undergo intensive dynamics. Tubulin dimers are incorporated at plus ends of spindle microtubules while they are removed from the minus ends, resulting in poleward movement. Such microtubule flux is regulated by the microtubule rescue factors CLASPs at kinetochores and depolymerizing protein Kif2a at the poles, along with other regulators of microtubule dynamics. How microtubule polymerization and depolymerization are coordinated remains unclear. Here we show that TPX2, a microtubule-bundling protein and activator of Aurora A, plays an important role. TPX2 was phosphorylated by Aurora A during mitosis. Its phospho-null mutant caused short metaphase spindles coupled with low microtubule flux rate. Interestingly, phosphorylation of TPX2 regulated its interaction with CLASP1 but not Kif2a. The effect of its mutant in shortening the spindle could be rescued by codepletion of CLASP1 and Kif2a that abolished microtubule flux. Together we propose that Aurora A-dependent TPX2 phosphorylation controls mitotic spindle length through regulating microtubule flux.


Subject(s)
Cell Cycle Proteins/metabolism , Kinesins/metabolism , Microtubule-Associated Proteins/metabolism , Microtubules/metabolism , Nuclear Proteins/metabolism , Spindle Apparatus/physiology , Animals , Aurora Kinase A/metabolism , Cell Line, Tumor , HeLa Cells , Humans , Metaphase/physiology , Mitosis/physiology , Phosphorylation , Protein Multimerization , Tubulin/metabolism , Xenopus
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