cellXpress: a fast and user-friendly software platform for profiling cellular phenotypes.

Background: High-throughput, image-based screens of cellular responses to genetic or chemical perturbations generate huge numbers of cell images. Automated analysis is required to quantify and compare the effects of these perturbations. However, few of the current freely-available bioimage analysis software tools are optimized for efficient handling of these images.

Mec1/Tel1-dependent phosphorylation of Slx4 stimulates Rad1-Rad10-dependent cleavage of non-homologous DNA tails.

Budding yeast Slx4 interacts with the Rad1-Rad10 endonuclease that is involved in nucleotide excision repair (NER), homologous recombination (HR) and single-strand annealing (SSA). We previously showed that Slx4 is dispensable for NER but is essential for SSA. Slx4 is phosphorylated by the Mec1 and Tel1 kinases after DNA damage on at least six Ser/Thr residues, and mutation of all six residues to Ala reduces the efficiency of SSA.

Coordination of structure-specific nucleases by human SLX4/BTBD12 is required for DNA repair.

Budding yeast Slx4 interacts with the structure-specific endonuclease Slx1 to ensure completion of ribosomal DNA replication. Slx4 also interacts with the Rad1-Rad10 endonuclease to control cleavage of 3' flaps during repair of double-strand breaks (DSBs). Here we describe the identification of human SLX4, a scaffold for DNA repair nucleases XPF-ERCC1, MUS81-EME1, and SLX1.

Phosphorylation of Slx4 by Mec1 and Tel1 regulates the single-strand annealing mode of DNA repair in budding yeast.

Budding yeast (Saccharomyces cerevisiae) Slx4 is essential for cell viability in the absence of the Sgs1 helicase and for recovery from DNA damage. Here we report that cells lacking Slx4 have difficulties in completing DNA synthesis during recovery from replisome stalling induced by the DNA alkylating agent methyl methanesulfonate (MMS). Although DNA synthesis restarts during recovery, cells are left with unreplicated gaps in the genome despite an increase in translesion synthesis.

Multiple approaches to study S. cerevisiae Rad9, a prototypical checkpoint protein.

The Saccharomyces cerevisiae RAD9 checkpoint gene is the prototypical checkpoint gene and is required for efficient checkpoint regulation in late G1, S, and at the G2/M cell cycle transition following DNA damage. Rad9 is required for the activation of Rad53 after damage and has been proposed to have roles in lesion recognition as well as DNA repair and the maintenance of genome stability. Here we describe methodology suitable for the study of G1, intra-S, and G2/M checkpoints in budding yeast, the analysis of Rad9/Rad53 phospho-forms, the biochemical analysis of Rad9 and Rad53, the fractionation of soluble and chromatin associated proteins, including Rad9, and the live cell imaging of GFP tagged Rad9.

Histone H2A phosphorylation and H3 methylation are required for a novel Rad9 DSB repair function following checkpoint activation.

In budding yeast, the Rad9 protein is an important player in the maintenance of genomic integrity and has a well-characterised role in DNA damage checkpoint activation. Recently, roles for different post-translational histone modifications in the DNA damage response, including H2A serine 129 phosphorylation and H3 lysine 79 methylation, have also been demonstrated. Here, we show that Rad9 recruitment to foci and bulk chromatin occurs specifically after ionising radiation treatment in G2 cells.

DNA repair: the importance of phosphorylating histone H2AX.

How phosphorylated histone H2AX, known as gamma-H2AX, functions in the cellular response to DNA double-strand breaks is the subject of intensive investigation. Recent research in yeast and mammalian cells shows that gamma-H2AX facilitates post-replicational DNA repair by recruiting cohesin, a protein complex that holds sister chromatids together.