Mastermind Mutations Generate a Unique Constellation of Midline Cells within the Drosophila CNS

Background: The Notch pathway functions repeatedly during the development of the central nervous system in metazoan organisms to control cell fate and regulate cell proliferation and asymmetric cell divisions. Within the Drosophila midline cell lineage, which bisects the two symmetrical halves of the central nervous system, Notch is required for initial cell specification and subsequent differentiation of many midline lineages. Methodology/Principal Findings: Here, we provide the first description of the role of the Notch co-factor, mastermind, in the central nervous system midline of Drosophila. Overall, zygotic mastermind mutations cause an increase in midline cell number and decrease in midline cell diversity. Compared to mutations in other components of the Notch signaling pathway, such as Notch itself and Delta, zygotic mutations in mastermind cause the production of a unique constellation of midline cell types. The major difference is that midline glia form normally in zygotic mastermind mutants, but not in Notch and Delta mutants. Moreover, during late embryogenesis, extra anterior midline glia survive in zygotic mastermind mutants compared to wild type embryos. Conclusions/Significance: This is an example of a mutation in a signaling pathway cofactor producing a distinct centra

The transcriptional coactivator MAML1 regulates p300 autoacetylation and HAT activity

MAML1 is a transcriptional coregulator originally identified as a Notch coactivator. MAML1 is also reported to interact with other coregulator proteins, such as CDK8 and p300, to modulate the activity of Notch. We, and others, previously showed that MAML1 recruits p300 to Notch-regulated genes through direct interactions with the DNA–CSL–Notch complex and p300. MAML1 interacts with the C/H3 domain of p300, and the p300–MAML1 complex specifically acetylates lysines of histone H3 and H4 tails in chromatin in vitro. In this report, we show that MAML1 potentiates p300 autoacetylation and p300 transcriptional activation. MAML1 directly enhances p300 HAT activity, and this coincides with the translocation of MAML1, p300 and acetylated histones to nuclear bodies

Guidelines for the use and interpretation of assays for monitoring autophagy (3rd edition)

In 2008 we published the first set of guidelines for standardizing research in autophagy. Since then, research on this topic has continued to accelerate, and many new scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Accordingly, it is important to update these guidelines for monitoring autophagy in different organisms. Various reviews have described the range of assays that have been used for this purpose. Nevertheless, there continues to be confusion regarding acceptable methods to measure autophagy, especially in multicellular eukaryotes. For example, a key point that needs to be emphasized is that there is a difference between measurements that monitor the numbers or volume of autophagic elements (e.g., autophagosomes or autolysosomes) at any stage of the autophagic process versus those that measure fl ux through the autophagy pathway (i.e., the complete process including the amount and rate of cargo sequestered and degraded). In particular, a block in macroautophagy that results in autophagosome accumulation must be differentiated from stimuli that increase autophagic activity, defi ned as increased autophagy induction coupled with increased delivery to, and degradation within, lysosomes (inmost higher eukaryotes and some protists such as Dictyostelium ) or the vacuole (in plants and fungi). In other words, it is especially important that investigators new to the fi eld understand that the appearance of more autophagosomes does not necessarily equate with more autophagy. In fact, in many cases, autophagosomes accumulate because of a block in trafficking to lysosomes without a concomitant change in autophagosome biogenesis, whereas an increase in autolysosomes may reflect a reduction in degradative activity. It is worth emphasizing here that lysosomal digestion is a stage of autophagy and evaluating its competence is a crucial part of the evaluation of autophagic flux, or complete autophagy. Here, we present a set of guidelines for the selection and interpretation of methods for use by investigators who aim to examine macroautophagy and related processes, as well as for reviewers who need to provide realistic and reasonable critiques of papers that are focused on these processes. These guidelines are not meant to be a formulaic set of rules, because the appropriate assays depend in part on the question being asked and the system being used. In addition, we emphasize that no individual assay is guaranteed to be the most appropriate one in every situation, and we strongly recommend the use of multiple assays to monitor autophagy. Along these lines, because of the potential for pleiotropic effects due to blocking autophagy through genetic manipulation it is imperative to delete or knock down more than one autophagy-related gene. In addition, some individual Atg proteins, or groups of proteins, are involved in other cellular pathways so not all Atg proteins can be used as a specific marker for an autophagic process. In these guidelines, we consider these various methods of assessing autophagy and what information can, or cannot, be obtained from them. Finally, by discussing the merits and limits of particular autophagy assays, we hope to encourage technical innovation in the field

Linking model systems to cancer therapeutics: the case of Mastermind

Genetics, and more recently genomics, reveal striking conservation in the fundamental signaling pathways that underlie normal and aberrant cell processes. Consequently, various genetic model organisms are now attracting the interest of biomedical scientists who are focused on therapeutic approaches to human disease. There are now several examples of studies in which Drosophila seems likely to facilitate advances in potential therapies, and a recent report has demonstrated the utility of the fly model for understanding and treating human disease. Basic developmental genetic information first obtained in Drosophila was used to design a therapeutic block to oncogenic Notch signaling that was associated with leukemia in mice. The story of Notch signaling in Drosophila demonstrates the potential for standard Drosophila molecular genetics in developing therapeutic strategies that are relevant to human disease

Genetic interactions of 30 chromatin protein encoding loci with <i>domino</i>.

<p>Wing mounts were prepared from the following Bloomington (BL) strains after crosses to <i>C96-domR</i>. Wings shown in panels C-F2 derived from crosses to TRiP strains. A. Wild Type wing BL3605 <i>w</i>^<i>1118</i>, B. Control wing <i>C96-domR/w</i>^<i>1118</i>, C. BL33981 <i>PCAF</i>, D. BL33962 <i>HP1c</i>, <i>E</i>. BL26234 <i>CC8</i>, <i>F</i>. BL31921 <i>JIGR1</i>, <i>G</i>. BL42514 <i>CC25</i>, <i>H</i>. BL31922 <i>CC20 I</i>. BL27085 <i>ERR</i>, <i>J</i>. BL33394 <i>RPS9</i>, <i>K</i>. BL32888 <i>CC24</i>, <i>L</i>. BL33361 <i>CC4</i>, <i>M</i>. BL33734 <i>CC7</i>, <i>N</i>. BL42491 <i>CC9</i>, <i>O</i>. BL26772 <i>CC15</i>, <i>P</i>. BL40853 <i>MAF-S</i>, <i>Q</i>. BL33974 <i>RYBP</i>, <i>R</i>. BL25993 <i>CC28</i>, <i>S</i>. BL29360 <i>CC32</i>, <i>T</i>. BL34580 <i>NUP50</i>, <i>U</i>. BL34069 <i>CAF1</i>, <i>V</i>. BL33043 <i>PCNA</i>, <i>W</i>. BL55250 <i>ASF1</i>, <i>X</i>. BL55314 <i>TOP1</i>, <i>Y</i>. BL33725 <i>RPD3</i>, <i>Z</i>. BL33666 <i>HEL25E</i>, <i>A2</i>. BL41937 <i>CC31</i>, <i>B2</i>. BL53697 <i>SIR2</i>, <i>C2</i>. BL34978 <i>TRIP1</i>, <i>D2</i>. BL26231 <i>CC34</i>, <i>E2</i>. BL31940 <i>CC30</i>, F2. BL31960 <i>DSP1</i></p