Biology subjects

van den Berg, A.

Publications and source records attributed to van den Berg, A..

3 recordsLinked to original sources

bioRxiv2021

Fluidic Circuit Board with Modular Sensor and Valves Enables Stand-Alone, Tubeless Microfluidic Flow Control in Organs-on-Chips

Organs-on-chips are a unique class of microfluidic in vitro cell culture models, in which the in vivo tissue microenvironment is mimicked. Unfortunately, its widespread use is hampered by their operation complexity and incompatibility with end-user research settings. To address these issues, many commercial and non-commercial platforms have been developed for semi-automated culture of organs-on-chips. However, these organ-on-chip culture platforms each represent a closed ecosystem, with very little opportunity to interchange and integrate components from different platforms or to develop new ones. The Translational Organ-on-Chip Platform (TOP) is a multi-institutional effort to develop an open platform for automated organ-on-chip culture and integration of components from various developers. Central to TOP is the fluidic circuit board (FCB), a microfluidic plate with the form factor of a typical well plate. The FCB enables microfluidic control of multiple components like sensors or organ-on-chip devices through an interface based on openly available standards. Here, we report an FCB to integrate commercial and in-house developed components forming a stand-alone flow control system for organs-on-chips. The control system is able to achieve constant and pulsatile flow recirculation through a connected organ-on-chip device. We demonstrate that this system is able to automatically perfuse a heart-on-chip device containing co-cultures of cardiac tissues derived from human pluripotent stem cell-derived cardiomyocytes and monolayers of endothelial cells for five days. Altogether, we conclude that open technology platforms allow the integration of components from different sources to form functional and fit-for-purpose organ-on-chip systems. We anticipate that open platforms will play a central role in catalysing and maturing further technological development of organ-on-chip culture systems.

bioengineering↗

bioRxiv2021

CRISPR/Cas9 screen for functional MYC binding sites reveals MYC-dependent vulnerabilities in K562 cells

The transcription factor MYC is a proto-oncogene with a well-documented essential role in the pathogenesis and maintenance of several types of cancer. MYC binds to specific E-box sequences in the genome to regulate gene expression in a cell type- and developmental stage-specific manner. To date, a comprehensive analysis of direct MYC targets with essential roles in different types of cancer is missing. To enable identification of functional MYC binding sites and corresponding target genes, we designed a CRISPR/Cas9 library to destroy E-box sequences in a genome-wide fashion. In parallel, we used the Brunello library to knockout protein-coding genes. We performed high-throughput screens with these libraries in four MYC-dependent cancer cell lines: K562, ST486, HepG2 and MCF7, which revealed several essential E-boxes and genes. Among them we pinpointed crucial known and novel MYC-regulated genes involved in pathways associated with cancer development. Extensive validation of our approach in K562 cells confirmed that E-box disruption affects MYC binding, target genes expression and cell proliferation. Our unique, well-validated tool opens new possibilities to gain novel insights into MYC-dependent vulnerabilities in cancer cells.

molecular biology↗

bioRxiv2019

RNA polymerases as moving barriers to condensin loop extrusion

To separate replicated sister chromatids during mitosis, eukaryotes and prokaryotes have structural maintenance of chromosome (SMC) condensin complexes that were recently shown to organize chromosomes by a process known as DNA loop extrusion. In rapidly dividing bacterial cells, the process of separating sister chromatids occurs concomitantly with ongoing transcription. How transcription interferes with the condensin loop extrusion process is largely unexplored, but recent experiments show that sites of high transcription may directionally affect condensin loop extrusion. We quantitatively investigate different mechanisms of interaction between condensin and elongating RNA polymerases (RNAP) and find that RNAPs are likely steric barriers that can push and interact with condensins. Supported by new Hi-C and ChIP-seq data for cells after transcription inhibition and RNAP degradation, we argue that translocating condensins must bypass transcribing RNAPs within ~2 seconds of an encounter at rRNA genes and within ~10 seconds at protein coding genes. Thus, while individual RNAPs have little effect on the progress of loop extrusion, long, highly transcribed operons can significantly impede the extrusion process. Our data and quantitative models further suggest that bacterial condensin loop extrusion occurs by two independent, uncoupled motor activities; the motors translocate on DNA in opposing directions and function together to enlarge chromosomal loops, each independently bypassing steric barriers in their path. Our study provides a quantitative link between transcription and 3D genome organization and proposes a mechanism of interactions between SMC complexes and elongating transcription machinery relevant from bacteria to higher eukaryotes.

biophysics↗