Centromeres are important structural constituents of chromosomes that ensure proper chromosome segregation during mitosis by providing defined sites for kinetochore attachment. In higher eukaryotes, centromeres have no specific DNA sequence and thus, they are rather determined through epigenetic mechanisms. A fundamental process in centromere establishment is the incorporation of the histone variant CENP-A into centromeric chromatin, which provides a binding platform for the other centromeric proteins. The Mis18 complex, and, in particular, its member M18BP1 was shown to be essential for both incorporation and maintenance of CENP-A.   Here we show that M18BP1 displays a cell cycle-regulated association with centromeric chromatin in mouse embryonic stem cells. M18BP1 is highly enriched at centromeric regions from late anaphase through to G1 phase. An interaction screen against 16 core centromeric proteins revealed a novel interaction of M18BP1 with CENP-C. We mapped the interaction domain in M18BP1 to a central region containing a conserved SANT domain and in CENP-C to the C-terminus. Knock-down of CENP-C leads to reduced M18BP1 association and lower CENP-A levels at centromeres, suggesting that CENP-C works as an important factor for centromeric M18BP1 recruitment and thus for maintaining centromeric CENP-A.

Dosage compensation in Drosophila involves a global activation of genes on the male X chromosome. The activating complex (MSL-DCC) consists of male-specific-lethal (MSL) proteins and two long, noncoding roX RNAs. The roX RNAs are essential for X-chromosomal targeting, but their contributions to MSL-DCC structure and function are enigmatic. Conceivably, the RNA helicase MLE, itself an MSL subunit, is actively involved in incorporating roX into functional DCC. We determined the secondary structure of roX2 and mapped specific interaction sites for MLE in vitro. Upon addition of ATP, MLE disrupted a functionally important stem loop in roX2. This RNA remodeling enhanced specific ATPdependent association of MSL2, the core subunit of the MSL-DCC, providing a link between roX and MSL subunits. Probing the conformation of roX in vivo revealed a remodeled stem loop in chromatin-bound roX2. The active remodeling of a stable secondary structure by MLE may constitute a ratelimiting step for MSL-DCC assembly.

We have identified a generalized arousal component in the behavior of mice. Analyzed by mathematical͞statistical approaches across experiments, investigators, and mouse populations, it accounts for about 1͞3 of the variance in arousal-related measures. Knockout of the gene coding for the classical estrogen receptor (ER-␣), a ligand-activated transcription factor, greatly reduced arousal responses. In contrast, disrupting the gene for a likely gene duplication product, ER-␤, did not have these effects. A combination of mathematical and genetic approaches to arousal in an experimentally tractable mammal opens up analysis of a CNS function of considerable theoretical and practical significance.estrogen ͉ motivation ͉ mice ͉ genomics ͉ estrogen receptor H ormonal, neural, and genetic mechanisms for simple sex behaviors in rats and mice have been worked out in some detail. Underlying all of these is sexual arousal. Concepts of arousal are essential for helping to explain broad classes of behavior, but they also have been murky and ill-defined. In humans, ''arousal'' is intuitively obvious, but what about in experimental animals?To justify mechanistic studies of arousal, we propose that neurophysiological and behavioral responsivity to external stimuli constitute elementary requirements for animal life. But whether there is a generalized arousal function has been hotly debated. Electrophysiological evidence from recordings across the cerebral cortex after manipulations of the brainstem said ''yes'' (1-3). However, some cognitive neuroscientists argued (4) that the concept of arousal has become hopelessly subdivided. A clear theoretical resolution can be found, for the first time, in an equation ʈ A ϭ F͑Kg⅐Ag ϩ Ks 1 ⅐As 1 ϩ Ks 2 ⅐As 2 ϩ Ks 3 ⅐As 3 ϩ ⅐ ⅐ ⅐ ϩ Ks n ⅐As n ͒,which combines both generalized (Ag) and various specific forms of arousal [As (1 to n) ] such as sex, hunger, fear, etc. (see also Fig. 1). Here, we show that principal components analysis (5) of published experimental data (6-9) shows quantitatively that generalized arousal influences the behavior of mice. New data from mice with the classical estrogen receptor (ER)-␣ or ER-␤ genes disrupted here illustrate how the gene for a particular nuclear receptor contributes to arousal. MethodsMice. The original five populations of ovariectomized female mice, the raw data from which have been reanalyzed by principal components analysis, were described in detail in refs. 6-9. Additional female mice were those in which the gene coding for ER-␣ had been functionally disrupted (n ϭ 8). These mice were originally obtained from the colony at the National Institute on Environmental Health Sciences (NIEHS, Research Triangle Park, NC) and had been backcrossed to a C57 background for at least eight generations. All their experimental procedures were run in parallel with WT littermate controls (n ϭ 9). Likewise, ER-␤ knockout females (n ϭ 10) were initially obtained from NIEHS, maintained and bred in our colony, and experimented in parallel with WT (n ϭ 10) littermate co...