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Genetic studies of memory formation in Drosophila have revealed that the formation of a protein synthesis-dependent long-term memory (LTM) requires multiple training sessions. LTM is blocked specifically by induced expression of a repressor isoform of the cAMP-responsive element-binding protein (CREB). Here, we report an enhancement of LTM formation after induced expression of an activator isoform of dCREB2. Maximum LTM is achieved after one training session, and its formation depends on phosphorylation of the activator transgene. A model of LTM formation based on differential regulation of CREB isoforms is proposed.

Periostin was originally identified as an osteoblast-specific factor and is highly expressed in the embryonic periosteum, cardiac valves, placenta, and periodontal ligament as well as in many adult cancerous tissues. To investigate its role during development, we generated mice that lack the periostin gene and replaced the translation start site and first exon with a lacZ reporter gene. Surprisingly, although periostin is widely expressed in many developing organs, periostin-deficient (peri lacZ ) embryos are grossly normal. Postnatally, however, ϳ14% of the nulls die before weaning and all of the remaining peri lacZ nulls are severely growth retarded. Skeletal analysis revealed that trabecular bone in adult homozygous skeletons was sparse, but overall bone growth was unaffected. Furthermore, by 3 months, the nulls develop an early-onset periodontal disease-like phenotype. Unexpectedly, these mice also show a severe incisor enamel defect, although there is no apparent change in ameloblast differentiation. Significantly, placing the peri lacZ nulls on a soft diet that alleviated mechanical strain on the periodontal ligament resulted in a partial rescue of both the enamel and periodontal disease-like phenotypes. Combined, these data suggest that a healthy periodontal ligament is required for normal amelogenesis and that periostin is critically required for maintenance of the integrity of the periodontal ligament in response to mechanical stresses.

The function of the epidermal growth factor receptor (EGFR) family member HER4 remains unclear because its activating ligand, heregulin, results in either proliferation or differentiation. This variable response may stem from the range of signals generated by HER4 homodimers versus heterodimeric complexes with other EGFR family members. The ratio of homo-and heterodimeric complexes may be influenced both by a cell's EGFR family member expression profile and by the ligand or even ligand isoform used. To define the role of HER4 in mediating antiproliferative and differentiation responses, human breast cancer cell lines were screened for responses to heregulin. Only cells that expressed HER4 exhibited heregulin-dependent antiproliferative responses. In-depth studies of one line, SUM44, demonstrated that the antiproliferative and differentiation responses correlated with HER4 activation and were abolished by stable expression of a kinase-inactive HER4. HB-EGF, a HER4-specific ligand in this EGFR-negative cell line, also induced an antiproliferative response. Moreover, introduction and stable expression of HER4 in HER4-negative SUM102 cells resulted in the acquisition of a heregulin-dependent antiproliferative response, associated with increases in markers of differentiation. The role of HER2 in these heregulin-dependent responses was examined through elimination of cell surface HER2 signaling by stable expression of a single-chain anti-HER2 antibody that sequestered HER2 in the endoplasmic reticulum. In the cell lines with either endogenously (SUM44) or exogenously (SUM102) expressed HER4, elimination of HER2 did not alter HER4-dependent decreases in cell growth. These results suggest that HER4 is both necessary and sufficient to trigger an antiproliferative response in human breast cancer cells.The epidermal growth factor receptor (EGFR) family has been implicated in breast cancer pathogenesis and progression (reviewed in references 13 and 39). Aberrant expression of at least two of the family members, EGFR and HER2, has been associated with poor prognosis and differential response to therapy (21,28,31,44). Recently, treatment targeted against HER2 has demonstrated clinical efficacy, emphasizing the importance of members of this receptor family in breast cancer prognosis and therapy (10).The EGFR family consists of four known members: EGFR (HER1, erbB-1), HER2 (erbB-2), HER3 (erbB-3), and HER4 (erbB-4) (reviewed in references 13, 34, and 39). The four receptors form homodimers or heterodimers upon activation by two sets of ligands, the EGF and heregulin/neuregulin families. There are several possible hetero-and homodimeric receptor combinations, which theoretically result in differential activation of multiple downstream signal transduction pathways. Additional heterogeneity results from varying phenotypic responses, depending on cell type and the duration or intensity of downstream signaling, determined in part by differences in ligand affinity, recycling, and intracellular environment, as well as other factors th...

Although mutated forms of ras are not associated with the majority of breast cancers (<5%), there is considerable experimental evidence that hyperactive Ras can promote breast cancer growth and development. Therefore, we determined whether Ras and Ras-responsive signaling pathways were activated persistently in nine widely studied human breast cancer cell lines. Although only two of the lines harbor mutationally activated ras, we found that five of nine breast cancer cell lines showed elevated active Ras-GTP levels that may be due, in part, to HER2 activation. Unexpectedly, activation of two key Ras effector pathways, the extracellular signal-regulated kinase (ERK) mitogen-activated protein kinase and phosphatidylinositol 3-kinase/AKT signaling pathways, was not always associated with Ras activation. Ras activation also did not correlate with invasion or the expression of proteins associated with tumor cell invasion (estrogen receptor ␣ and cyclooxygenase 2). We then examined the role of Ras signaling in mediating resistance to matrix deprivationinduced apoptosis (anoikis). Surprisingly, we found that ERK and phosphatidylinositol 3-kinase/AKT activation did not have significant roles in conferring anoikis resistance. Taken together, these observations show that Ras signaling exhibits significant cell context variations and that other effector pathways may be important for Ras-mediated oncogenesis, as well as for anoikis resistance, in breast cancer. Additionally, because ERK and AKT activation are not strictly associated with Ras activation, pharmacological inhibitors of these two signaling pathways may not be the best approach for inhibition of aberrant Ras function in breast cancer treatment.

We report the role of dCREB2, the Drosophila homolog of CREB/CREM, in circadian rhythms. dCREB2 activity cycles with a 24 hr rhythm in flies, both in a light:dark cycle and in constant darkness. A mutation in dCREB2 shortens circadian locomotor rhythm in flies and dampens the oscillation of period, a known clock gene. Cycling dCREB2 activity is abolished in a period mutant, indicating that dCREB2 and Period affect each other and suggesting that the two genes participate in the same regulatory feedback loop. We propose that dCREB2 supports cycling of the Period/Timeless oscillator. These findings support CREB's role in mediating adaptive behavioral responses to a variey of environmental stimuli (stress, growth factors, drug addiction, circadian rhythms, and memory formation) in mammals and long-term memory formation and circadian rhythms in Drosophila.

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