CDC7 kinase promotes MRE11 fork processing, modulating fork speed and chromosomal breakage.

The CDC7 kinase is essential for the activation of DNA replication origins and has been implicated in the replication stress response. Using a highly specific chemical inhibitor and a chemical genetic approach, we now show that CDC7 activity is required to coordinate multiple MRE11-dependent processes occurring at replication forks, independently from its role in origin firing. CDC7 localizes at replication forks and, similarly to MRE11, mediates active slowing of fork progression upon mild topoisomerase inhibition.

Transport of soluble proteins through the Golgi occurs by diffusion via continuities across cisternae.

The mechanism of transport through the Golgi complex is not completely understood, insofar as no single transport mechanism appears to account for all of the observations. Here, we compare the transport of soluble secretory proteins (albumin and α1-antitrypsin) with that of supramolecular cargoes (e.g.

Silencing of mammalian Sar1 isoforms reveals COPII-independent protein sorting and transport.

The Sar1 GTPase coordinates the assembly of coat protein complex-II (COPII) at specific sites of the endoplasmic reticulum (ER). COPII is required for ER-to-Golgi transport, as it provides a structural and functional framework to ship out protein cargoes produced in the ER. To investigate the requirement of COPII-mediated transport in mammalian cells, we used small interfering RNA (siRNA)-mediated depletion of Sar1A and Sar1B.

ADP-ribose polymer depletion leads to nuclear Ctcf re-localization and chromatin rearrangement(1).

Ctcf (CCCTC-binding factor) directly induces Parp [poly(ADP-ribose) polymerase] 1 activity and its PARylation [poly(ADPribosyl)ation] in the absence of DNA damage. Ctcf, in turn, is a substrate for this post-synthetic modification and as such it is covalently and non-covalently modified by PARs (ADP-ribose polymers). Moreover, PARylation is able to protect certain DNA regions bound by Ctcf from DNA methylation.

Secretory traffic triggers the formation of tubular continuities across Golgi sub-compartments.

The organization of secretory traffic remains unclear, mainly because of the complex structure and dynamics of the secretory pathway. We have thus studied a simplified system, a single synchronized traffic wave crossing an individual Golgi stack, using electron tomography. Endoplasmic-reticulum-to-Golgi carriers join the stack by fusing with cis cisternae and induce the formation of intercisternal tubules, through which they redistribute their contents throughout the stack.

Golgi enzymes are enriched in perforated zones of golgi cisternae but are depleted in COPI vesicles.

In the most widely accepted version of the cisternal maturation/progression model of intra-Golgi transport, the polarity of the Golgi complex is maintained by retrograde transport of Golgi enzymes in COPI-coated vesicles. By analyzing enzyme localization in relation to the three-dimensional ultrastructure of the Golgi complex, we now observe that Golgi enzymes are depleted in COPI-coated buds and 50- to 60-nm COPI-dependent vesicles in a variety of different cell types. Instead, we find that Golgi enzymes are concentrated in the perforated zones of cisternal rims both in vivo and in a cell-free system.