The pseudogene is required to safeguard germ cells against reprogramming.

We recently identified FAcilitates Chromatin Transcription (FACT) as a reprogramming barrier of transcription factor (TF) mediated conversion of germ cells into neurons in . FACT is a conserved heterodimer consisting of SPT16 and SSRP1 in mammals. Duplication events during evolution in generated two SSRP1 homologs named HMG-3 and HMG-4, while SPT-16 is the only homolog of SPT16.

The CONJUDOR pipeline for multiplexed knockdown of gene pairs identifies RBBP-5 as a germ cell reprogramming barrier in C. elegans.

Multiple gene activities control complex biological processes such as cell fate specification during development and cellular reprogramming. Investigating the manifold gene functions in biological systems requires also simultaneous depletion of two or more gene activities. RNA interference-mediated knockdown (RNAi) is commonly used in Caenorhabditis elegans to assess essential genes, which otherwise lead to lethality or developmental arrest upon full knockout.

Strategies for in vivo reprogramming.

Reprogramming has the potential to provide specific cell types for regenerative medicine applications aiming at replacing tissues that have been lost or damaged due to degenerative diseases and injury. In this review we discuss the latest strategies and advances of in vivo reprogramming to convert cell identities in living organisms, including reprogramming induced by transcription factors (TFs) and CRISPR/dCas9 synthetic TFs, as well as by cell fusion and small molecules. We also provide a brief recap of reprogramming barriers, the effect of senescence on reprogramming efficiency, and strategies to deliver reprogramming factors in vivo.

MRG-1/MRG15 Is a Barrier for Germ Cell to Neuron Reprogramming in .

Chromatin regulators play important roles in the safeguarding of cell identities by opposing the induction of ectopic cell fates and, thereby, preventing forced conversion of cell identities by reprogramming approaches. Our knowledge of chromatin regulators acting as reprogramming barriers in living organisms needs improvement as most studies use tissue culture. We used as an gene discovery model and automated solid-phase RNA interference screening, by which we identified 10 chromatin-regulating factors that protect cells against ectopic fate induction.

FACT Sets a Barrier for Cell Fate Reprogramming in Caenorhabditis elegans and Human Cells.

The chromatin regulator FACT (facilitates chromatin transcription) is essential for ensuring stable gene expression by promoting transcription. In a genetic screen using Caenorhabditis elegans, we identified that FACT maintains cell identities and acts as a barrier for transcription factor-mediated cell fate reprogramming. Strikingly, FACT's role as a barrier to cell fate conversion is conserved in humans as we show that FACT depletion enhances reprogramming of fibroblasts.

ECM modulated early kidney development in embryonic organ culture.

The use of exogenous signals is gaining importance in renal regenerative therapies. We wanted to explore the role of extracellular matrix (ECM) constituents on renal structure formation during renal organogenesis. We used a recently established organ culture setup to expose embryonic kidney rudiments directly to a large set of surface-immobilized or dissolved ECM molecules and growth factors.

Dewaxed ECM: A simple method for analyzing cell behaviour on decellularized extracellular matrices.

Decellularization techniques have been used on a wide variety of tissues to create cell-seedable scaffolds for tissue engineering. Finding a suitable decellularization protocol for a certain type of tissue can be laborious, especially when organ perfusion devices are needed. In this study, we report a quick and simple method for comparing decellularization protocols combining the use of paraffin slices and two-dimensional cell cultures.

A novel, low-volume method for organ culture of embryonic kidneys that allows development of cortico-medullary anatomical organization.

Here, we present a novel method for culturing kidneys in low volumes of medium that offers more organotypic development compared to conventional methods. Organ culture is a powerful technique for studying renal development. It recapitulates many aspects of early development very well, but the established techniques have some disadvantages: in particular, they require relatively large volumes (1-3 mls) of culture medium, which can make high-throughput screens expensive, they require porous (filter) substrates which are difficult to modify chemically, and the organs produced do not achieve good cortico-medullary zonation.