scholarly journals Active establishment of centromeric CENP-A chromatin by RSF complex

2009 ◽  
Vol 185 (3) ◽  
pp. 397-407 ◽  
Author(s):  
Marinela Perpelescu ◽  
Naohito Nozaki ◽  
Chikashi Obuse ◽  
Hua Yang ◽  
Kinya Yoda

Centromeres are chromosomal structures required for equal DNA segregation to daughter cells, comprising specialized nucleosomes containing centromere protein A (CENP-A) histone, which provide the basis for centromeric chromatin assembly. Discovery of centromere protein components is progressing, but knowledge related to their establishment and maintenance remains limited. Previously, using anti-CENP-A native chromatin immunoprecipitation, we isolated the interphase–centromere complex (ICEN). Among ICEN components, subunits of the remodeling and spacing factor (RSF) complex, Rsf-1 and SNF2h proteins, were found. This paper describes the relationship of the RSF complex to centromere structure and function, demonstrating its requirement for maintenance of CENP-A at the centromeric core chromatin in HeLa cells. The RSF complex interacted with CENP-A chromatin in mid-G1. Rsf-1 depletion induced loss of centromeric CENP-A, and purified RSF complex reconstituted and spaced CENP-A nucleosomes in vitro. From these data, we propose the RSF complex as a new factor actively supporting the assembly of CENP-A chromatin.

2010 ◽  
Vol 298 (2) ◽  
pp. G248-G254 ◽  
Author(s):  
Bradley L. Urquhart ◽  
Jamie C. Gregor ◽  
Nilesh Chande ◽  
Michael J. Knauer ◽  
Rommel G. Tirona ◽  
...  

Folic acid is a vitamin essential for thymidylate and purine synthesis. The human proton-coupled folate transporter (hPCFT) has recently been identified as a pH-dependent folic acid transporter, and mutations in this transporter have been linked to hereditary folic acid malabsorption. In this study, we assessed hPCFT-mediated transport activity in vitro, intersubject variability of intestinal expression in relation to blood folates, and the relationship of proton-pump inhibitor (PPI) therapy on hPCFT expression in vivo. We created a Madin-Darby canine kidney strain II (MDCKII) cell line stably expressing hPCFT to evaluate its drug substrates and inhibitors. Intestinal pinch biopsies (duodenum, ileum, colon) were collected from patients undergoing routine endoscopy procedures, and expressed levels of hPCFT were determined by RT-PCR. When assessed using MDCKII-hPCFT cells, folic acid and methotrexate were found to be high-affinity hPCFT substrates. Sulfasalazine and pyrimethamine were noted to inhibit hPCFT activity with Ki values of 42.3 and 161.7 μmol/l, respectively. hPCFT was localized to the brush-border membrane of enterocytes with highest expression in the duodenum and reduced levels in the ileum and colon. When we assessed hPCFT expression in a subset of patients who were receiving PPI therapy, a near 50% reduction in duodenal hPCFT mRNA expression was noted. These results suggest that hPCFT transporter activity can be modulated by many drugs in clinical use, and expression of this transporter in the gastrointestinal tract is higher in the duodenum than more distal sites (duodenum > ileum > colon). Importantly, we note that PPI drug use appears to be associated with reduced hPCFT expression in vivo.


2001 ◽  
Vol 155 (7) ◽  
pp. 1147-1158 ◽  
Author(s):  
Samantha G. Zeitlin ◽  
Richard D. Shelby ◽  
Kevin F. Sullivan

Aurora B is a mitotic protein kinase that phosphorylates histone H3, behaves as a chromosomal passenger protein, and functions in cytokinesis. We investigated a role for Aurora B with respect to human centromere protein A (CENP-A), a centromeric histone H3 homologue. Aurora B concentrates at centromeres in early G2, associates with histone H3 and centromeres at the times when histone H3 and CENP-A are phosphorylated, and phosphorylates histone H3 and CENP-A in vitro at a similar target serine residue. Dominant negative phosphorylation site mutants of CENP-A result in a delay at the terminal stage of cytokinesis (cell separation). The only molecular defects detected in analysis of 22 chromosomal, spindle, and regulatory proteins were disruptions in localization of inner centromere protein (INCENP), Aurora B, and a putative partner phosphatase, PP1γ1. Our data support a model where CENP-A phosphorylation is involved in regulating Aurora B, INCENP, and PP1γ1 targeting within the cell. These experiments identify an unexpected role for the kinetochore in regulation of cytokinesis.


1971 ◽  
Vol 49 (1) ◽  
pp. 131-NP ◽  
Author(s):  
G. P. VINSON ◽  
J. G. PHILLIPSt ◽  
I. CHESTER JONES ◽  
W. N. TSANG

SUMMARY The relationship of structure and function in the adrenal gland of the possum Trichosurus vulpecula, has been studied using in-vitro incubation techniques. It was shown that both 17α-hydroxycorticosteroids and 17-deoxycorticosteroids were produced from radioactive pregnenolone and progesterone, and that these transformations occurred both in the definitive cortex as well as in a special zone of hypertrophied tissue found only in the adult female. In support of earlier findings, it was also shown that the adrenal cortex of the possum has a remarkable capacity to produce C19 steroids (including androstenedione and testosterone) from the radioactive precursors. While most of the transformations occurred with equal efficiency in both types of tissue, the reduction of androstenedione to testosterone seemed to take place more readily in the special hypertrophied zone of the adult female. In studies in vivo, it was found that levels of testosterone in the peripheral blood of the adult female possum were extremely high compared with man. Variations in testosterone levels were not apparently correlated with the stage of the oestrous cycle. The possible pathways by which the adrenal products are synthesized, and their physiological implications are discussed.


2000 ◽  
Vol 97 (3) ◽  
pp. 1148-1153 ◽  
Author(s):  
E. V. Howman ◽  
K. J. Fowler ◽  
A. J. Newson ◽  
S. Redward ◽  
A. C. MacDonald ◽  
...  

2000 ◽  
Vol 97 (13) ◽  
pp. 7266-7271 ◽  
Author(s):  
K. Yoda ◽  
S. Ando ◽  
S. Morishita ◽  
K. Houmura ◽  
K. Hashimoto ◽  
...  

Author(s):  
J.R. Pfeiffer ◽  
J.C. Seagrave ◽  
C. Wofsy ◽  
J.M. Oliver

In RBL-2H3 rat leukemic mast cells, crosslinking IgE-receptor complexes with anti-IgE antibody leads to degranulation. Receptor crosslinking also stimulates the redistribution of receptors on the cell surface, a process that can be observed by labeling the anti-IgE with 15 nm protein A-gold particles as described in Stump et al. (1989), followed by back-scattered electron imaging (BEI) in the scanning electron microscope. We report that anti-IgE binding stimulates the redistribution of IgE-receptor complexes at 37“C from a dispersed topography (singlets and doublets; S/D) to distributions dominated sequentially by short chains, small clusters and large aggregates of crosslinked receptors. These patterns can be observed (Figure 1), quantified (Figure 2) and analyzed statistically. Cells incubated with 1 μg/ml anti-IgE, a concentration that stimulates maximum net secretion, redistribute receptors as far as chains and small clusters during a 15 min incubation period. At 3 and 10 μg/ml anti-IgE, net secretion is reduced and the majority of receptors redistribute rapidly into clusters and large aggregates.


Blood ◽  
1948 ◽  
Vol 3 (7) ◽  
pp. 729-754 ◽  
Author(s):  
WILLIAM N. VALENTINE ◽  
CHARLES G. CRADDOCK ◽  
JOHN S. LAWRENCE

Abstract The hormonal control through the hypophyseo-adrenal cortical system of lymphoid tissue structure and function is an important concept. We cannot at the present time regard that the concept is established fact. Final judgment must await additional work and the clarification of some of the inconsistencies which appear to exist. It seems reasonable that lymphoid tissue is one of the end organs of adrenal cortical hormone and that it may perhaps play a role in the response of the organism to stress. It seems quite clear that the sugar hormone of the adrenal cortex is capable of producing structural alterations in lymphoid tissue. Change in thoracic duct lymphocyte numbers as a result of augmentation in the amount of available adrenal cortical hormone is at present controversial. Experiments in this laboratory have failed to demonstrate it. The production of lymphopenia, at least in some species and possibly in man, by increasing available sugar hormone is supported by some evidence. The exact mechanism of production of lymphopenia is open to question, its relationship to changes in lymphoid tissue structure being one of inference. The converse situation—absolute lympocytosis resulting from deprivation of adrenal cortical hormone—is the subject of controversial reports. At best, it must be admitted that relatively slight alterations from the accepted normal range of lymphocyte values occur in the adrenal insufficient organism. Changes in plasma gamma globulins and antibody titers associated with changes in the amount of available cortical hormone are reported. It should be clarified whether such changes have necessarily resulted from lymphocyte dissolution or are related to other of the variegated actions of adrenal cortical hormone. The relationship of adrenal cortical hormone to lymphoid tissue and lymphocytes and the relationship of the latter to the response of the organism to stress must indeed be complex. It is reasonably well established that the life span of the lymphocyte is very short indeed1,58,22 and each lymphocyte presumably liberates its metabolically important contents within a few hours at the most. If stress continues for any period of time, as often it does, it is difficult to visualize the wisdom of interfering with the production of metabolically vital substances in order to secure the transient benefits of lymphoid tissue dissolution. It is also somewhat difficult to regard as proved that the various changes reported after hormone augmentation or deprivation necessarily represent the normal mechanism by which these factors are regulated and kept within physiologic limits. More investigations are required to answer such questions and to further elucidate the interrelationship of the adrenal cortex and lymphoid tissues.


1980 ◽  
Vol 9 (4) ◽  
pp. 179-183 ◽  
Author(s):  
H L Stark ◽  
A Al-Haboubi

The relationships of width, thickness, volume and load to extension for human skin in vitro are reported. The specimens tested exhibited a low stiffness phase followed by a high stiffness phase. Volume rose than fell back to the initial volume at approximately the end of the low stiffness phase, and continued on falling to a final reduction of about 20 per cent at failure. Width decreased throughout, showing a maximum rate of reduction at approximately the end of the low stiffness phase. Thickness increased at a rate which also was maximum at the end of the low stiffness phase. The specimens used were long compared with their width and thickness thus offering no constraint to lateral contraction. An interpretation of this data in respect of the behaviour of the collagen fibre matrix is postulated.


1977 ◽  
Vol 43 (5) ◽  
pp. 839-843 ◽  
Author(s):  
J. A. Severson ◽  
R. D. Fell ◽  
J. G. Tuig ◽  
D. R. Griffith

Plasma corticosterone concentrations and in vitro adrenal secretion of corticosterone were determined in exercise-trained rats. Rats, 100, 200, and 300 days of age, were trained for a 10-wk period by treadmill running. Following the training program, rats were subjected to an acute bout of swimming. Acute swimming elevated plasma corticosterone concentrations in all age groups. At 170 days of age, the plasma corticosterone concentration following swimming was higher in exercise-trained rats than in controls. The opposite was true of acutely swum rats at 270 and 370 days of age. Acute swimming elevated the in vitro adrenal gland response to adrenocorticotropic hormone stimulation in control rats at all ages and in trained rats at 170 days of age. The in vivo relationship of epinephrine and the pituitary adrenal system is suggested as a mechanism which could have caused this response. The relationship of secretion rates to plasma corticosterone concentrations indicated that extra-adrenal mechanisms, such as decreased turnover, were also responsible for the elevated plasma corticosterone levels observed in response to acute swimming.


1964 ◽  
Vol 6 (2) ◽  
pp. 169-178 ◽  
Author(s):  
R. V. Large

1. Thirty Suffolk × Half bred lambs were slaughtered at the following ages: two twin lambs at birth and two singles and two twins at 1, 2, 3, 4, 6, 8 and 16 weeks of age.2. The following weights were recorded: live-weight immediately before slaughter; and carcass, head, skin, feet, alimentary tract, heart, liver, kidneys, lungs and trachea, and blood immediately afterwards.3. The alimentary tract was emptied and weighed in four separate parts; reticulo-rumen, omasum-abomasum, small intestine, large intestine.4. The volumes of the reticulo-rumen and the omasum-abomasum were measured by immersing in water and filling the organs with water to 2 cm. pressure.5. The in vitro digestive efficiency of rumen liquor from lambs of 1, 2, 3 and 4 weeks of age was assessed.6. Empty body weight was considered to be valuable in comparing animals of different ages or from different feeding regimes or at different times of the year because variations in gut ‘fill’ were eliminated.7. There were no differences between singles and twins in the relationship of the fresh weights of the parts of the body to empty body weight, except that development of the liver and the blood was rather slower for singles.8. Little evidence was found of a difference in rate of development of the alimentary tract between singles an d twins, although the log an d square root transformation suggested a possible difference in reticulo-rumen size in favour of twins, significant at the 5% level.


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