Histones are dynamically modified during chromatin assembly, as specific transcriptional patterns are established, and during mitosis and development. Modifications include acetylation, phosphorylation, ubiquitination, methylation, and ADP-ribosylation, but the biological significance of each of these is not well understood. For example, distinct acetylation patterns correlate with nucleosome formation and with transcriptionally activated or silenced chromatin, yet mutations in genes encoding several yeast histone acetyltransferase (HAT) activities result in either no cellular phenotype or only modest growth defects. Here we report characterization of ESA1, an essential gene that is a member of the MYST family that includes two yeast silencing genes, human genes associated with leukemia and with the human immunodeficiency virus type 1 Tat protein, and Drosophila mof, a gene essential for male dosage compensation. Esa1p acetylates histones in a pattern distinct from those of other yeast enzymes, and temperature-sensitive mutant alleles abolish enzymatic activity in vitro and result in partial loss of an acetylated isoform of histone H4 in vivo. Strains carrying these mutations are also blocked in the cell cycle such that at restrictive temperatures, esa1 mutants succeed in replicating their DNA but fail to proceed normally through mitosis and cytokinesis. Recent studies show that Esa1p enhances transcription in vitro and thus may modulate expression of genes important for cell cycle control. These observations therefore link an essential HAT activity to cell cycle progression, potentially through discrete transcriptional regulatory events.

Post-translational acetylation of histone H4 N-terminal tail in chromatin has been associated with several nuclear processes including transcription. We report the purification and characterization of a native multisubunit complex (NuA4) from yeast that acetylates nucleosomal histone H4. NuA4 has an apparent molecular mass of 1.3 MDa. All four conserved lysines of histone H4 can be acetylated by NuA4. We have identified the catalytic subunit of the complex as the product of ESA1, an essential gene required for cell cycle progression in yeast. Antibodies against Esa1p specifically immunoprecipitate NuA4 activity whereas the complex purified from a temperature-sensitive esa1 mutant loses its acetyltransferase activity at the restrictive temperature. Additionally, we have identified another subunit of the complex as the product of TRA1, an ATM-related essential gene homologous to human TRRAP, an essential cofactor for c-Myc-and E2F-mediated oncogenic transformation. Finally, the ability of NuA4 to stimulate GAL4-VP16-driven transcription from chromatin templates in vitro is also lost in the temperature-sensitive esa1 mutant. The function of the essential Esa1 protein as the HAT subunit of NuA4 and the presence of Tra1p, a putative transcription activator-interacting subunit, supports an essential link between nuclear H4 acetylation, transcriptional regulation and cell cycle control.

Silencing is an epigenetic form of transcriptional regulation whereby genes are heritably, but not necessarily permanently, inactivated. We have identified the Saccharomyces cerevisiae genes SAS2 and SAS3 through a screen for enhancers of sir1 epigenetic silencing defects. SAS2, SAS3 and a Schizosaccharomyces pombe homologue are closely related to several human genes, including one associated with acute myeloid leukaemia arising from the recurrent translocation t(8;16)(p11;p13) and one implicated in HIV-1 Tat interactions. All of these genes encode proteins with an atypical zinc finger and well-conserved similarities to acetyltransferases. Sequence similarities and yeast mutant phenotypes suggest that SAS-like genes function in transcriptional regulation and cell-cycle exit and reveal novel connections between transcriptional silencing and human disease.

Objective-Despite the existence of several follow-up studies of children with ADHD followed up into adulthood, there is limited information on whether patterns of persistence and remission in ADHD can be predicted over the long-term. The main aim of this study was to evaluate predictors of persistence of ADHD in a large sample of boys with and without ADHD followed prospectively for 11 years into young adulthood.Method-Subjects were Caucasian, non-Hispanic boys with (N=110) and without (N=105) ADHD who were 6 to 17 years old at the baseline assessment (mean age 11 years) and 15 to 31 years old at the follow-up assessment (mean age 22 years). Subjects were comprehensively and blindly assessed with structured diagnostic interviews and assessments of cognitive, social, school, and family functioning.Results-At the 11-year follow-up, 78% of children with ADHD continued to have a full (35%) or a partial persistence (subsyndromal (22%), impaired functioning (15%), or remitted but treated (6%)). Predictors of persistence were severe impairment of ADHD, psychiatric comorbidity, and exposure to maternal psychopathology at baseline.Conclusions-These findings prospectively confirm that persistence of ADHD over the long term is predictable from psychosocial adversity and psychiatric comorbidity ascertained 11 years earlier.

The Mac-2 lectin (carbohydrate binding protein 35) is a soluble, 32-to 35-kDa phosphoprotein that binds galactose-containing glycoconjugates. We report here that the colonic epithelium is a major site of Mac-2 expression in vivo based on immunohistochemistry of human tissue specimens. In this epithelium, proliferating cells at the base of the crypts do not express Mac-2 but its expression increases with differentiation along the crypt-to-surface axis. Mac-2 expression is concentrated in the nuclei of these differentiated epithelial cells.The progression from normal mucosa to adenoma to carcinoma is associated with significant changes in Mac-2 nuclear localization and expression. In all adenomas (9/9) and carcinomas (13/13) examined, Mac-2 was not present in the nucleus but was localized in the cytoplasm. Sequencing of Mac-2 cDNAs from normal mucosa and carcinoma revealed no specific mutations that could account for this loss of nuclear localization. We also observed a 5-to 10-fold decrease in Mac-2 mRNA levels in cancer compared to normal mucosa as well as a significant reduction in the amount of Mac-2 protein expressed. These observations suggest that Mac-2 exclusion from the nucleus and its decreased expression may be related to the neoplastic progression of colon cancer.The Mac-2 lectin is a soluble, 32-to 35-kDa phosphoprotein that binds galactose-containing glycoconjugates (1-5). Molecular cloning studies revealed that Mac-2 is identical to carbohydrate binding protein 35 (CBP 35), a lectin characterized initially in 3T3 fibroblasts (3,4,6). It is also identical to IgE binding protein (7) and a 32-kDa tumor-associated lectin (8, 9). The COOH-terminal domain of Mac-2, which contains a conserved carbohydrate binding motif, is homologous to a related family of 14-kDa galactose-specific lectins (reviewed in ref. 5). Collectively, Mac-2 and the 14-kDa lectins are referred to as S-lectins because their carbohydrate binding function was thought to be thiol-dependent (10), an assumption that has recently been questioned (2).The biological functions of Mac-2 remain elusive. Its putative role in cell adhesion has not been substantiated (11). Mac-2 may be associated with cell growth and differentiation because it is found in the nucleus of 3T3 fibroblasts and its nuclear localization may be related to the proliferative state of these cells (5). Also, the expression of Mac-2 occurs as a function of macrophage differentiation (12). These reports are consistent with the recent finding that a homologue of the 14-kDa S-lectin is cytostatic for embryonic fibroblasts (13). Other in vitro studies have suggested that the expression of Mac-2 increases in oncogenically transformed and metastatic cells (14-16).Progress in elucidating the function of Mac-2 has been hampered, in part, by a lack of data on its expression in vivo.In the present study, we found that Mac-2 is largely an epithelial-specific lectin with high expression seen in the colonic epithelium. The progression from normal colonic mucosa to adenoma to carcin...

Objective Deficient emotional self regulation (DESR) is characterized by deficits in self-regulating the physiological arousal caused by strong emotions. We examined whether a unique profile of the Child Behavior Check List (CBCL) would help identify DESR in children with Attention- Deficit/ Hyperactivity Disorder (ADHD). Methods Subjects were 197 children with and 224 without ADHD. We defined DESR if a child had an aggregate cut-off score of > 180 but < 210 on the Anxiety/Depression, Aggression, and Attention scales of the CBCL (CBCL-DESR). This profile was selected because of 1) its conceptual congruence with the clinical concept of DESR and 2) because its extreme (>210) form had been previously associated with severe forms of mood and behavioral dysregulation in children with ADHD. All subjects were comprehensively assessed with structured diagnostic interviews and a wide range of functional measures. Results Forty four percent of children with ADHD had a positive CBCL- DESR profile vs. 2% of controls (p<0.001). The CBCL-DESR profile was associated with elevated rates of anxiety and disruptive behavior disorders, as well as significantly more impairments in emotional and interpersonal functioning. Conclusions The CBCL-DESR profile helped identify a subgroup of ADHD children with a psychopathological and functional profile consistent with the clinical concept of DESR.

Accurate segregation of chromosomes is critical to ensure that each daughter cell receives the full genetic complement. Maintenance of cohesion between sister chromatids, especially at centromeres, is required to segregate chromosomes precisely during mitosis and meiosis. The Drosophila protein MEI-S332, the founding member of a conserved protein family, is essential in meiosis for maintaining cohesion at centromeres until sister chromatids separate at the metaphase II/anaphase II transition. MEI-S332 localizes onto centromeres in prometaphase of mitosis or meiosis I, remaining until sister chromatids segregate. We elucidated a mechanism for controlling release of MEI-S332 from centromeres via phosphorylation by POLO kinase. We demonstrate that POLO antagonizes MEI-S332 cohesive function and that full POLO activity is needed to remove MEI-S332 from centromeres, yet this delocalization is not required for sister chromatid separation. POLO phosphorylates MEI-S332 in vitro, POLO and MEI-S332 bind each other, and mutation of POLO binding sites prevents MEI-S332 dissociation from centromeres.

The integrin ␣ 6 ␤ 4 , a receptor for members of the laminin family of basement membrane components, contributes to the function of epithelial cells and their oncogenically transformed derivatives. In our efforts to study ␣ 6 ␤ 4 -mediated functions in more detail and to assess the contribution of the ␤ 4 cytoplasmic domain in such functions, we identified a rectal carcinoma cell line that lacks expression of the ␤ 4 integrin subunit. This cell line, termed RKO, expresses ␣ 6 ␤ 1 but not ␣ 6 ␤ 4 , and it interacts with laminin-1 less avidly than similar cell lines that express ␣ 6 ␤ 4 . We expressed a full-length ␤ 4 cDNA, as well as a mutant cDNA that lacks the ␤ 4 cytoplasmic domain, in RKO cells and isolated stable subclones of these transfectants. In this study, we report that subclones that expressed the full-length ␤ 4 cDNA in association with endogenous ␣6 exhibited partial G 1 arrest and apoptosis, properties that were not evident in RKO cells transfected with either the cytoplasmic domain mutant or the expression vector alone. In an effort to define a mechanism for these observed changes in growth, we observed that expression of the ␣ 6 ␤ 4 integrin induced expression of the p21 (WAF1; CiP1) protein, an inhibitor of cyclin-dependent kinases. These data suggest that the ␤ 4 integrin cytoplasmic domain is linked to a signaling pathway involved in cell cycle regulation in the ␤ 4 transfected RKO cells.The integrin ␣ 6 ␤ 4 is a receptor for members of the laminin family of basement membrane components. Initial studies established that ␣ 6 ␤ 4 is a receptor for laminin-1 (1-3), and subsequent work has shown that it also functions as a receptor for other laminin isoforms (4, 5). In its capacity as a laminin receptor, ␣ 6 ␤ 4 is involved in the formation and maintenance of hemidesmosomes (6 -8) and in the dynamic adhesion and migration of carcinoma cells (2, 3). Most likely, other functions of epithelial and carcinoma cells are dependent upon ␣ 6 ␤ 4 because it plays such a pivotal role in mediating their interactions with laminin matrices. It is widely assumed that the unusually large and structurally unique cytoplasmic domain of the ␤ 4 integrin subunit associates with cytoskeletal and signaling molecules and that such associations provide the basis for the distinct functions associated with ␣ 6 ␤ 4 (5, 8, 9).In our efforts to study ␣ 6 ␤ 4 -mediated functions in more detail and to assess the contribution of the ␤ 4 cytoplasmic domain in such functions, we identified a rectal carcinoma cell line that lacks expression of the ␤ 4 integrin subunit. This cell line, termed RKO, expresses ␣ 6 ␤ 1 but not ␣ 6 ␤ 4 (2, 3). In this study, we report that RKO transfectants, which expressed the fulllength ␤ 4 cDNA in association with endogenous ␣ 6 exhibited G 1 arrest and a basal rate of apoptosis, properties that were not evident in RKO cells transfected with either a ␤ 4 cytoplasmic domain mutant or the expression vector alone. In an effort to define a mechanism for these observed changes in growth, we observed that e...