circRNAs (zirkuläre RNAs) in neuralen Zellen

N. Rajewsky Lab

Systems Biology of Gene Regulatory Elements

Profile

Research Interests and Approaches

The Rajewsky lab studies how RNA regulates gene expression. A past focus of the group was on mechanisms and functions of microRNAs. More recently, we have started to study the function of RNA binding proteins and long noncoding RNAs, including circular RNAs (circRNAs). We are using various model systems.

For understanding the role of noncoding RNA in (a) early development (b) stem cell biology and regeneration (c) function of neurons we use C. elegans, planaria, and mice, respectively. In collaboration with colleagues at the Charité, we are also analyzing human brain samples. Recently, we have established human brain organoids in the lab.

We integrate experimental (biochemistry, molecular biology) and computational (bioinformatics, physics, statistics) methods. We have developed several widely used methods (for example, recently, “MirDeep” (Friedlander et al. Nature Biotechnology 2008, Friedlaender et al. NAR 2012); miRNA target identification by chimera analyses (Grosswendt et al. Mol. Cell 2014); fixation method for droplet based single cell sequencing (Alles et al. BMC Biology 2017); “DistMap” for mapping sequenced single cells to correct positions within complex tissues (Karaiskos et al. Science 2017)).  A few years ago we implemented and optimized “drop-seq” and are using it in many projects to sequence RNA from single cells in high throughput (e.g. Karaiskos et al. Science 2017). We also develop methods to detect, quantify, image, and knockdown/knock out circRNAs (Memczak et al. Nature 2013, Rybak-Wolf et al. Mol. Cell 2014, Piwecka et al. Science 2017).

Jobs

We are always looking for skilled people!

Whether your weapon of choice is pipette or keyboard (or both): if you have the impression that this lab is doing interesting research that you would like to contribute to, do not hesitate to contact us.

We are an international and interdisciplinary team (biologists, biochemists, bioinformaticians, physicists, ...) and communication is key. If you are able to (and enjoy) discussing your work with people from other disciplines this lab may be just right!

How to get to our lab

House 87: Timoféeff-Ressovsky-House (Genomcentrum)

The Rajewsky lab is located on the MDC (Max Delbrück Center) campus, building 87 (Timofeeff Ressovsky House, or “Genome Center“).

The MDC campus itself is in the north east of Berlin. The neighborhood is called "Berlin Buch" and is essentially a small town.

Our address

Robert Rössle Str 10
13092 Berlin-Buch
Phone +49 30 9406 2999 (Alex Tschernycheff, Secretary)
Fax +49 30 9406 3868

The MDC provides information about how to get to the MDC but in our PDF we provide some more practical help.

Google Maps

Team

Lab retreat to MeckPom 2017

As in previous years the group went to the idyllic Gutshaus Rensow in Mecklenburg-Vorpommern in September to enjoy stimulating scientific discussions in a beautiful setting.

Jonathan Alles

Ph.D. Student

Contact information:

  • E-mail: jonathan dot alles at mdc-berlin dot de
  • Phone: +49 30 9406 2996

Start date: June 2017

Funding: NYU-MDC Exchange Program

"I am interested in developing and applying experimental and computational approaches for exploring transcript isoform expression at single cell resolution"

Salah Ayoub

Technician

Contact information:

E-mail: salah dot ayoub at mdc-berlin dot de

Phone: +49 30 9406 2973

Start date: December 2008

Funding: MDC

"I am mainly involved in Planaria projects, implementing diverse techniques (RNAi, injections, FACS, etc.). My tasks include lab organization; monitoring and establishment of lab processes, implementation and optimization of protocols, data generation and verification, import/export of biological materials, purchasing of supplies, contact to commercial representatives, management of equipment, next-generation sequencing (NGS) technologies, maintenance of animals (S. mediterranea, C. elegans), and running internal databases (development and administration)."

Anastasiya Boltengagen

Technician

Contact information:

  • E-mail: anastasiya dot boltengagen at mdc-berlin dot de
  • Phone: +49 30 9406 2972

Start date: November 2014

Funding: BIH

"research statement to follow"

Asija Diag

Ph.D. Student

Contact information:

  • E-mail: asija dot diag at mdc-berlin dot de
  • Phone: +49 30 9406 2972

Start date: September 2015

Funding: DFG/CSB

"A spatially resolved RNA map for the Caenorhabditis elegans germline"

Jonathan Fröhlich

Ph.D. Student

Contact information:

  • E-mail: jonathan dot froehlich at mdc-berlin dot de
  • Phone: +49 30 9406 2996
  • Start date: January 2014

Funding: MDC

"I am investigating the biological role of regulatory elements in RNA.  For this, I work with the roundworm C. elegans and establish methods for efficient genome editing."

Petar Glazar

Ph.D. Student

Contact information:

  • E-mail: petar dot glazar at mdc-berlin dot de
  • Phone: +49 30 9406 3044

Start date: November 2013

Funding: DEEP

"My research focuses on understanding the biogenesis and functions of circRNAs, particularly in early neurogenesis. I am also developing methods and tools for circRNA analysis and quantification from RNA-seq data."

Margareta Herzog

Lab Manager

Contact information:

  • E-mail: margareta dot herzog at mdc-berlin dot de
  • Phone: +49 30 9406 2972 (lab), -2973 (lab)

Start date: March 2011

Funding: MDC

Cledi Cerda Jara

Ph.D. Student

Contact information:

  • E-mail: cledi dot cerdajara at mdc-berlin dot de
  • Phone: +49 30 9406 2972

Start date: February 2016

Funding: MDC Graduate School

"Studying the function of circRNAs in neurons, and the mechanisms underlying their role"  

Nikos Karaiskos

Postdoctoral Researcher

Contact information:

  • E-mail: nikolaos dot karaiskos at mdc-berlin dot de
  • Phone: +49 30 9406 2993

Start date: February 2015

Funding: DFG

"My research focus is the biology at the single-cell level and in particular the study of cell-cell interactions, by using computational and mathematical methods."

Seung Joon Kim

Ph.D. Student

Contact information:

  • E-mail: seungjoon dot kim at mdc-berlin dot de

Start date: July 2017

Funding: EU/ITN

"I am interested in analyzing single cell RNA-seq data to identify mechanisms involved in pathology of neurological disorders by computational approaches"

David Koppstein

Postdoctoral Researcher

Contact information:

  • E-mail: david dot koppstein at mdc-berlin dot de
  • Phone: +49 30 9406 2991

Start date: June 2018

Funding: MDC

Ivano Legnini

Postdoctoral Reseacher

Contact information:

  • E-mail: ivano dot legnini at mdc-berlin dot de
  • Phone: +49 30 9406 2989

Start date: March 2017

Funding: EMBO fellowship

"I'm currently studying RNA translation and localization in neurons, by applying various sequencing-based technologies and trying to develop new methods for addressing these fundamental problems."  

Haiyue Liu

Ph.D. Student

Contact information:

  • E-mail: haiyue dot liu at mdc-berlin dot de
  • Phone: +49 30 9406 2989

Start date: September 2016

Funding: CSC scholarship

"I'm interested in developing methods for pathway analysis of single cell RNA-seq data."

Gwendolin Matz

Technical Assistant

Contact information:

  • E-mail: gwendolin dot matz at mdc-berlin dot de
  • Phone: +49 30 9406 2972

Start date: March 2017

Funding: DZHK

Aristotelis Misios

Ph.D. Student

Contact information:

  • E-mail: aristotelis dot misios at mdc-berlin dot de
  • Phone: +49 30 9406 2992

Start date: November 2016

Funding: EU/ITN

Panagiotis Papavasileiou

Ph.D. Student

Contact information:

  • E-mail: panagiotis dot papavasileiou at mdc-berlin dot de
  • Phone: +49 30 9406 2991

Start date: August 2013

Funding: MDC NYU Exchange programme

"In order to improve the understanding of biogenesis and function of circular RNAs, I am working on their screening in a variety of organisms and cell cultures. In particular, I'm interested in the impact of circular RNAs in development and differentiation of D. melanogaster."  

Monika Piwecka

Postdoctoral Researcher

Contact information:

  • E-mail: monika dot piwecka at mdc-berlin dot de
  • Phone: +49 30 9406 2972

Start date: February 2015

Funding: MDC

"I’m interested in non-coding RNAs and their roles in posttranscriptional gene regulatory networks in neurons and brain pathologies. Particular focus on circRNAs, their biology and functions, and miRNA."

Mireya Plass Pórtulas

Postdoctoral Researcher

Contact information:

  • E-mail: mireya dot plassportulas at mdc-berlin dot de
  • Phone: +49 30 9406 4248

Start date: January 2016

Funding: DZHK

"My main research interest is to study the mechanisms that regulate gene expression among cells with the same genetic background. In particular, I use single cell transcriptomics to understand how gene expression changes during differentiation and cell cycle progression."  

Samantha Praktiknjo

Postdoctoral Researcher

Contact information:

  • E-mail: samantha dot praktiknjo at mdc-berlin dot de
  • Phone: +49 30 9406 3027

Start date: February 2016

Funding: MDC

"Characterization of tumor heterogeneity by single-cell sequencing."

Agnieszka Rybak-Wolf

Postdoctoral Researcher

Contact information:

  • E-mail: agnieszka dot rybak at mdc-berlin dot de
  • Phone: +49 30 9406 2972

Start date: October 2009

Funding: BIH

"I am interested in the circular RNAs expression, biogenesis and function in mammalian brain. A particular focus is on investigating their possible role in the control of neuronal identity and regulation of  adult neurogenesis."

Marcel Schilling

Ph.D. Student

Contact information:

  • E-mail: marcel dot schilling  at mdc-berlin dot de
  • Phone: +49 30 9406 4267

Start date: November 2013

Funding: MDC"I am interested in the post-transcriptional code, with a focus on small RNAs. In silico prediction of miRNA genes and target sites have been challenging tasks for computational biologists over the last years. I am trying to identify small regulatory RNAs and the mechanisms underlying their function based on sequencing data. Using this knowledge I want to help understanding disease pathogenesis, particularly the transition from acute to chronic inflammation."

Tamas Sztanka-Toth

Ph.D. Student

Contact information:

  • E-mail: tamasryszard dot sztanka-toth at mdc-berlin dot de
  • Phone: +49 30 9406 4267

Start date: September 2017

Funding: EU/ITN

Christin Sünkel

Ph.D. Student

Contact information:

  • E-mail: christin dot stottmeister at mdc-berlin dot de
  • Phone: +49 30 9406 2973

Start date: April 2014

Funding: Boehringer Ingelheim

"My aim is to elucidate the role of circular RNAs in neuronal differentiation and disease. Therefore I use human cell culture in gain- and loss-of-function studies, as well as pulldown approaches."

Kathrin Theil

Ph.D. Student

Contact information:

  • E-mail: kathrin dot theil at mdc-berlin dot de
  • Phone: +49 30 9406 2972

Start date: September 2010

Funding: HFSP

"I am interested in the role of RNA-binding proteins (RBPs) in posttranscriptional gene regulation. My research centres on i) the mechanisms underlying translational control by RBPs and on ii) competition effects in posttranscriptional gene regulation. To address my questions, I employ cell culture and transgenics in C. elegans combined with high-throughput methods like mass spectrometry and transcriptome-wide methods like PAR-CLIP."

Alex Tschernycheff

Administration

Contact information:

  • E-mail: tschernycheff at mdc-berlin dot de
  • Phone: +49 30 9406 2999
  • Fax: +49 30 9406 3068

Start date: May 2006

Funding: MDC

Marco Uhrig

Ph.D. Student

Contact information:

  • E-mail: marco dot uhrig at mdc-berlin dot de
  • Phone: +49 30 9406 2996
  • Fax: +49 30 9406 3068

Start date: February 2015

Funding: MDC Graduate School

"My PhD project is dealing with coding and regulatory functions of small open reading frames (ORFs) that are evolutionary conserved in animals. In order to elucidate their uncharacterized properties and the functional role of encoded micropeptides, I am using a combination of computational and experimental approaches including genome engineering via CRISPR/Cas9 in both cell lines and model organisms."  

Vera Zywitza

Ph.D. Student

Contact information:

  • E-mail: vera dot zywitza at mdc-berlin dot de
  • Phone: +49 30 9406 2973
  • Fax: +49 30 9406 3068

Start date: April 2014

Funding: MDC

"My research centers on the question how stem cells are regulated post transcriptionally. Therefore, I study RNA-protein interactions in the stem cell model system S. mediterranea."

Associated PI

Christine Kocks

Associate PI

Contact information:

  • E-mail: christine dot kocks at mdc-berlin dot de
  • Phone: +49 30 9406 3537

Start date: January 2012

Funding: MDC

"I am interested in systems biology approaches to study gene regulation in tissue development, repair and regeneration. I will use C. elegans and Planaria as model systems and try to apply or refine genetic techniques for the microscopic tracking of individual RNA-protein complexes. In close collaboration with the Rajewsky lab I hope to uncover evolutionarily conserved mechanisms that will further our understanding of human stem cells and cancer."

Former Lab Members

Sebastian Memczak

was a Ph.D student and Postdoc in our lab

will be working as postdoc at Salk Institute in California

Benedikt Obermayer

was a Postdoctoral Researcher in our lab

is now working at the BIH

Current address

Berlin

Andrei Filipchyk

was a Ph.D student in our lab

is now working as a scientific researcher at the Forschungszentrum Juelich/current address Juelich

Marta Rodriguez-Orejuela

was a Ph.D student in our lab

Current address:

Berlin

Luisa Schreyer

was a Technician in our lab

Jordi Solana Garcia

was a Postdoc in our lab

is now a Group Leader at Oxford Brooks University

Current address:

Oxford, UK

Filippos Klironomos

was a Postdoc in our lab

is now a Senior Scientist at Charité - University Medicine Berlin

Current address:

Berlin

Catherine Adamidi

was a Senior Scientist in our lab and after that in Tom Tuschl's lab. Catherine is back in Berlin now.

Current address:

Berlin

Kevin Chen

was a Postdoc in our lab

is now Assistant Professor with his own group

Current address:

Rutgers University, New York, USA

Antigoni Elefsinioti

was a Postdoc in our lab

is now a Postdoc at Bayer Health Care

Current address:

Bayer Health Care, Berlin, Germany

Marc Friedländer

was a Ph.D. student and for a time a Postdoc in our lab and subsequently an EMBO PostDoc with Roderic Guigo

is now Assistant Professor in RNA Biology at Stockholm University

Current Address:

Stockholm University, Science for Life Laboratory, Solna, Sweden

Stefanie Grosswendt

was a Ph.D. Student in our lab and worked shortly at the BMBF. She is now...

Current address:

BMBF, Bundesministerium für Bildung und Forschung, Berlin, Germany

Dominic Grün

was a Postdoc in our lab and a Postdoc in Alexander van Oudenaarden's lab

is now an Independent Group Leader at the MPI in Freiburg

Current address:

Max Planck Institut, Freiburg, Germany

Andranik Ivanov

was a Ph.D. student in our lab and is now a Postdoc, working for Dieter Beule at the Bioinformatics Core Unit at the BIH

Current address:

Berlin Institut of Health (BIH), Berlin, Germany

Anna-Carina Jungkamp

was a Ph.D. student in our lab, then joined McKinsey & Company, and after that Nils Bluethgen's lab as a Postdoc. Works now at the BMBF.

Current Address:

BMBF - Bundesministerium fuer Bildung und Forschung

Marvin Jens

was a Ph.D. student and postdoc in our lab and is now working at the MIT in Boston in the lab of Christopher Burge

Current address:

MIT Boston, USA

Toshiaki Kogame

was a Ph.D. student

is now a doctor at Kyoto Medical University

Current Address:

Kyoto Medical University, Kyoto, Japan

Azra Krek

was a Ph.D. student

is now a staff scientist at Sloan-Kettering, New York

Current Address:

Memorial Sloan-Kettering Cancer Center, New York, USA

Svetlana Lebedeva

was a Ph.D. Student, worked several years as a postdoc with Rene Ketting and recently joined the lab of Uwe Ohler at BIMSB/MDC

Current address:

IMB Mainz, Germany

Jonas Maaskola

was a Ph.D in our lab

is now a Postdoc at KTH Royal Institute of Technology in Stockholm

Current address:

KTH Royal Institute of Technology, Stockholm, Sweden

Sebastian Mackowiak

was a Ph.D in our lab

is now a Postdoc in Marc Friedlaender's lab in Stockholm

Current address:

Stockholm University, Science for Life Laboratory, Solna, Sweden

Pinar Önal

was a Ph.D in our lab

is now a Postdoc with Steve Small

Current address:

New York University, New York USA

Lena von Oertzen

was a TA in our lab and is now working at the FMP in Berlin-Buch

Current address:

FMP

Marlon Stoeckius

was a Ph.D. student and susequently a Postdoc in our lab, left for the USA to work with Antonio Girald. He is now working at the New York Genome Center.

Current Address:

New York Genome Center, NYU, New York, USA

Nadine Thierfelder

was a Ph.D. Student

is now an account executive at Hill+Knowlton Strategies GmbH

Current address

Hill+Knowlton Strategies GmbH, Frankfurt a.M., Germany 

Francesca Torti

was a Ph.D. Student

Current address

tbc

Ulrike Ziebold

was working as a Postdoc in our lab

Current address

tbc

Minnie Zhou Fang

was a PostDoc in our lab and a PostDoc in Ria Baumgrass' lab. She moved back to China and is now a Principle Scientist at Eli Lilly Research and Development in Shanghai

Current address:

Eli Lilly Research and Development, Shanghai, China

Research

Examples of research projects

Example 1: Gene expression in the fly embryo at single cell resolution

The lab has a long standing interest in the role of RNA in developmental biology. We established and optimized “drop-seq” in our lab (Alles et al. 2017). This microfluidics based method allows to rapidly capture mRNAs within thousands of individual cells in an hour on a desktop setup. After amplification and sequencing, based on barcode information individual reads can be assigned to individual molecules in individual cells.

In collaboration with the Robert Zinzen lab, we applied this method to the dissociated fly embryo at stage 6. Each of the 6,000 cells of this embryo is distinct as it will give rise to a different part of the fly body. Therefore, it is important to know the molecular makeup of each individual cell.

However, after sequencing individual cells, the problem is to identify the spatial position of this cell within the embryo. We designed a new method that solves this problem (“Distmap”) (Karaiskos et al., Science 2017).

As a result, we quantified gene expression for the vast majority of the embryo cells at a resolution of ~8,000 genes per cell. These data allowed us to create a “virtual embryo” where researchers can perform, online, virtual in situ hybridization for thousands of genes (www.devtex.org).

We identified dozens of transcription factors and lncRNAs as likely novel regulatory factors in this system, and discovered that Hippo signaling is active in the embryo and probably responsible for inducing the exit of patches of cells from synchronized cell divisions, thus explaining observations that have been made decades ago.

Gene expression in the fly embryo at single cell resolution (Karaiskos et al. Science 2017)

Example 2: Human brain organoids

Very recently, we have established human brain organoids in the lab (Agnieszka Rybak-Wolf, unpublished). This system allows us to study, at single cell and subcellular level, functions of regulatory RNA. This system also allows us to study human brain diseases in individual patients since we can derive organoids by reprogramming cells from an individual into pluripotent stem cells. Using CRISP-CAS we can also edit the genome within organoids.

Example 3: Regulation by RNA

We are studying regulation by RNA in solid tumors, muscle cells in human patients, and cardio-vascular systems in collaboration with many colleagues.

Example 4: Circular RNAs

In 2012/2013, we discovered that animals express a large number of different circular RNAs (circRNAs) in the cytoplasm of cells (Memczak et al. Nature 2013). This observation was also published independently by the Brown and Sharpless labs.

However, we also showed that circRNA expression is tissue and developmental stage specific and identified a circRNA (CDR1as) that we proposed regulates gene expression by binding large numbers of a conserved miRNA (miR-7) (Memczak et al. Nature 2013; back-to-back with the Kjems lab).

Our paper also contained several methods for detecting, imaging, and quantifying circRNA expression. We thus proposed that circRNAs can have regulatory roles. This is interesting since circRNAs are highly stable and therefore may perform specific tasks.

In a serious of follow up papers, we (a) showed that circRNAs are usually produced by “backsplicing” (Ashwal-Fluss et al. Mol Cell 2014) (b) published a widely used public circRNA database where we curate published circRNAs (Glazar et al. RNA 2015) (c) showed that thousands of circRNAs are readily detected in human blood and may serve as biomarkers (d) discovered that a few hundred circRNAs are highly expressed in the mammalian brain and that this expression is well conserved, developmental stage- and tissue specific, and enriched in synaptosomes (Rybak-Wolf et al. Mol. Cell 2015).

Finally, we removed a the CDR1as locus from the mouse genome, showed that the expression of miRNAs that directly interact with CDR1as is specifically altered in the brain of KO mice, and found a behavior phenotype (“pre-pulse inhibition deficit”) that is indicative of neuropsychiatric disorders as well as an electrophysiological synaptic phenotype (Piwecka et al. Science 2017). We believe that these results argue that we discovered a regulatory layer of circRNAs and interacting molecules that have regulatory function in the mammalian brain.

Example 5: Competition effects in post-transcriptional gene regulation

We wrote an “Analysis Review” about functions of competition effects in post-transcriptional gene regulation. In this review, we also proposed and tested one of the first quantitative models. (Jens & Rajewsky, Nature Reviews Genetics 2014).

Model organisms

We study posttranscriptional gene regulation in cell culture and two famous model systems: The nematode Caenorhabditis elegans (and its relative Caenorhabditis briggsae) and Schmidtea mediterranea.

C. elegans

The early embryo and the germline of C.elegans are highly regulated on the posttranscriptional level. For example, while PolII activity starts at the 4-cell stage, the earlier divisions already differentiate the future germline from the soma and set the body axes, without any transcription at all. [Stitzel and Seydoux 2007, Yamamoto, I.; Kosinski, M. E. & Greenstein, D. (2006)]

© Marlon Stoeckius

Background information

An ideal system to explore very early developmental processes

Since its establishment as a model organism, Caenorhabditis elegans has been an invaluable tool for biological research. An immense spectrum of questions can be addressed using this small nematode, making it one of the most versatile and exciting model organisms.The worm is also the organism in which miRNAs have been first described.

The worm is an ideal system to explore the very early developmental processes because of its rapid and invariant development and its ease of RNAi and transgenic techniques. Studies in the model organism C. elegans have yielded most of what is known today about metazoan development. The first embryonic division in C. elegans has been intensely studied since it is used as an in vivo system for detailed studies of metazoan cell division.

Very early development in the worm (and most other species) is mainly controlled by maternal gene products which are loaded into the oocyte during oogenesis. After fertilization the C. elegans embryo undergoes a series of stereotyped asymmetric cleavages that spatially segregate these maternal factors, such as transcription factors and coding and noncoding RNAs. After a set of divisions the embryo activates its own transcription. This so called maternal to zygotic transition is a universal process in animal development: The embryo overtakes the control and thus no longer solely relies on maternally provided transcripts.

We are investigating the function of small RNAs during very early development of C. elegans.

S. mediterranea

The fresh water flatworm S. mediterranea has been described already more than 200 years ago. Like many flatworms, it exhibits an astounding capacity to regenerate after injuries: a single animal, cut into dozens of pieces, may give rise to dozens of fully regenerated, clonal animals.

Also, unlike another famous regeneration model organism, the radial-symmetric Hydra, the bilateral planarians are complex, with an organized nervous system, eyes and other organs.
The young Charles Darwin collected some land planarians during his voyage with the HMS Beagle (1831 to 1836). He notes:

"Having cut one of them transversely into two nearly equal parts, in the course of a fortnight both had the shape of perfect animals. I had, however, so divided the body, that one of the halves contained both the inferior orifices, and the other, in consequence, none. In the course of twenty-five days from the operation, the more perfect half could not have been distinguished from any other specimen.
The other had increased much in size; and towards its posterior end, a clear space was formed in the parenchymatous mass, in which a rudimentary cup-shaped mouth could clearly be distinguished; on the under surface, however, no corresponding slit was yet open. If the increased heat of the weather, as we approached the equator, had not destroyed all the individuals, there can be no doubt that this last step would have completed in structure. Although so well known an experiment, it was interesting to watch the gradual production of every essential organ, out of the simple extremity of another animal."
["The Voyage of the Beagle" pp.26]

 

© Catherine Adamidi, MDC

 

Background information

 

The Rajewsky lab established planaria as a model system in the lab. These freshwater flatworms are famous for their almost unlimited ability to regenerate any tissue via pluripotent, adult stem cells. We are studying the role of small RNAs in planarian regeneration.

Publications

News

Data, Software & Resources

Discrover

A motif discovery method to find binding sites of nucleic acid binding proteins

Find further details on the Discrover website.

The corresponding publication is:

"Binding site discovery from nucleic acid sequences by discriminative learning of hidden Markov models"
Jonas Maaskola and Nikolaus Rajewsky
Nucleic Acid Research, 42(21):12995-13011, Dec 2014. doi:10.1093/nar/gku1083

Planaria

Useful resources for the planarian research community

Gene models

isotigs_all_full_length.fa

The transcripts sequences were obtained by genome independent de novo transcriptome assembly (Adamidi et al.(2011)) combined with genome based transcript predictions inferred with CUFFLINKS (Trapnell et al. (2010)). For details, see Adamidi et al. (2011) and Oenal et al. (2012).

Genomic transcript coordinates

isotigs_all_unique.gtf

If a transcripts maps to multiple genomic contigs, only the longest matching region is reported.

Expression data

Oenal_et_al_2012_mRNA_and_protein_expression_data.csv

The file contains transcript and protein expression data for all genes. If the protein expression was quantified for peptides in different translated frames (due to frame-shifting errors in the genome independent de novo annotation) the table contains multiple entries for the respective gene.

See the README for details on the table format.

eFACS

A method to collect staged C. elegans embryos by fluorescence-activated cell sorting

We devised a method to collect staged C. elegans embryos by fluorescence-activated cell sorting (eFACS). A single eFACS run routinely yields tens of thousands of almost perfectly staged one-cell stage embryos.

In principle, eFACS can be used to extract large samples of embryos enriched in any desired embryonic stage. Thus, eFACS opens the door to apply many modern high-throughput technologies to assay embryonic developmental stage specific gene expression in C. elegans. Several of such investigations are already ongoing in the lab.

iPAR-CLIP

A PAR-CLIP protocol for living C.elegans animals

Our lab established a PAR-CLIP protocol for living C.elegans animals, called iPAR-CLIP. This allows to study RNA-binding protein interactions in vivo. Here, we list our published work, using this method. All sequencing data of published experiments are available through GEO.

In Vivo and Transcriptome-wide Identification of RNA Binding Protein Target Sites

Anna-Carina Jungkamp , Marlon Stoeckius , Desirea Mecenas , Dominic Grün , Guido Mastrobuoni , Stefan Kempa , Nikolaus Rajewsky

Mol Cell. 2011 Dec 9;44(5):828-40. doi: 10.1016/j.molcel.2011.11.009.

PDF GEO raw data

Unambiguous identification of miRNA:target site interactions by different types of ligation reactions.

Grosswendt S, Filipchyk A, Manzano M, Klironomos F, Schilling M, Herzog M, Gottwein E, Rajewsky N.

Mol Cell. 2014 Jun 19;54(6):1042-54. doi: 10.1016/j.molcel.2014.03.049. Epub 2014 May 22.

PDF GEO raw data

A variety of dicer substrates in human and C. elegans.

Rybak-Wolf A, Jens M, Murakawa Y, Herzog M, Landthaler M, Rajewsky N

Cell. 2014 Nov 20;159(5):1153-67. doi: 10.1016/j.cell.2014.10.040.

PDF GEO raw data

A note on RNA thio labeling

The above papers utilized 4-thiouridine for RNA-labeling and crosslinking. We have also attempted to use 4-thiouracil as an alternative to 4-thiouridine for iPAR-CLIP in C.elegans. The raw data can be made available upon request by E-mail to Nikolaus Rajewsky.

PicTar

An algorithm for the identification of microRNA targets

PicTar is a project of the Rajewsky lab at NYU's Center for Comparative Functional Genomics and the MDC, Berlin.

The PicTar website provides details (3' UTR alignments with predicted sites, links to various public databases etc) regarding:

  1. microRNA target predictions in vertebrates (Krek et al, Nature Genetics 37:495-500 (2005))
  2. microRNA target predictions in seven Drosophila species (Grün et al, PLoS Comp. Biol. 1:e13 (2005))
  3. microRNA targets in three nematode species (Lall et al, Current Biology 16, 1-12 (2006))
  4. human microRNA targets that are not conserved but co-expressed (i.e. the microRNA and mRNA are expressed in the same tissue) (Chen and Rajewsky, Nat Genet 38, 1452-1456 (2006))
  5. UPDATED PICTAR PREDICTIONS 2012: doRiNA: a database of RNA interactions in post-transcriptional regulation (Anders et al, Nucleic Acids Res. 2012 Jan;40(Database issue):D180-6. Epub 2011 Nov 15.)

Go to PicTar

Bulk Downloads

PicTar miRNA target site predictions for the hg17, mm7, dm2 and ce2 genomes can be obtained from the UCSC genome browser via the 'tables' feature.
New predictions for hg18, mm9 and ce6 are available on our own UCSC mirror at http://dorina.mdc-berlin.de

For your convenience, doRiNA offers a link for downloading PicTar target site predictions: simply run a doRiNA query for all miRNA targets, setting the search options to 100% under step 3, and follow the link on the top of the results page.

AlignmentBuilder

To build contiguous alignments for each 3' UTR or other types of mRNAs (5' UTRs, CDS, ..) from the MultiZ multiple alignments at the UCSC Genome Database you can use the scripts from AlignmentBuilder.

Notes

  • The format of the gene annotation table available from UCSC changes occasionally. Check that the strand is given as "+" or "-", and not just "+" or blank. In the latter case you need to insert the missing "-"
  • The program also requires that the gene annotation table have exactly 10 fields - no "bin" field at the beginning and no extra fields at the end. Currently, there is a short script, transform_annotation.py that removes the first field and the extra fields at the end, but watch this in the future in case the format changes again.
  • There is an off-by-one error with the multiZ alignments file. You need to add 1 to the second field of that file or the alignments will be off by one.
  • Do not use the characters "." or "-" in the gene name because this will cause errors.

miReduce

 

miReduce correlates the logarithm of expression fold changes of a set of genes with the motif content of the regulatory sequences of these genes. In particular, it correlates the genome wide mRNA log fold changes in experiments involving the knockout/knockdown/overexpression of one or more miRNAs with the motif content of the 3'UTRs. The model is based on linear regression, and is intended for genome-wide studies.

The code is written as a Perl script for Linux. The details on how to use it and how to interpret the results are in the README file, as well as the literature where more details can be found.

Download the miReduce package

To unpack, do the 'tar -xzf miReduce.tgz'. This will create a directory called miReduce.

If you decide to try and use miReduce, please read the README file carefully, as this is a somewhat rough implementation that we still decided to release due to high demand for this kind of tool.

RNA competition

 

We here provide data and code to reproduce the analyses and plots from the following publication:

“Competition between target sites of regulators shapes post-transcriptional gene regulation”.
Nature Reviews Genetics (Analysis) 2014 (in press, doi:10.1038/nrg3853) Jens M, Rajewsky N.

Download

The tar archive contains python scripts that work with python 2.7, numpy, and matplotlib. Please see the contained README and LICENSE for details.

You can download the tarball here.

miRDeep2

Discovering known and novel miRNAs from deep sequencing data

Developed by Sebastian Mackowiak, Marc Friedländer and Nikolaus Rajewsky
Systems Biology group at the Max Delbrück Center, Berlin-Buch
April 20th, 2011

miRDeep2 is a completely overhauled tool which discovers microRNA genes by analyzing sequenced RNAs. The tool reports known and hundreds of novel microRNAs with high accuracy in seven species representing the major animal clades. The low consumption of time and memory combined with user-friendly interactive graphic output makes miRDeep2 accessible for straightforward application in current reasearch.

Download: miRDeep2.0.0.8 (Latest Update: May 2016) contains the core algorithm script and all auxiliary scripts.

The package is freely available for non-commercial purposes. For commercial purposes, please contact us.

For general comments or questions on commercial use, contact: rajewsky at mdc-berlin dot de.

Disclaimer

The miRDeep2 software package is provided as is without any guarantees or warranty for correctness. The authors are not responsible for any damage or loss of any kind caused by the use or misuse of the scripts included in the software package. The authors are not under obligation to provide support, service, corrections, or upgrades to the package.

miRDeep

Developed by Marc Friedländer and Nikolaus Rajewsky
Systems Biology group at the Max Delbrück Center, Berlin-Buch
April 4th, 2008

The miRDeep package was developed to discover active known or novel miRNAs from deep sequencing data (Solexa/Illumina, 454, ...). The package consists of everything you need to analyze your own deep sequencing data after removal of ligation adapters: a number of scripts to preprocess the mapped data, and the core miRDeep algorithm that will analyze and score these data.

The package is freely available for non-commercial purposes. For commercial purposes, please contact us.

If you use miRDeep in your scientific research, please cite us:

Friedländer, M.R., Chen, W., Adamidi, C., Maaskola, J., Einspanier, R., Knespel, S., Rajewsky, N. 'Discovering microRNAs from deep sequencing data using miRDeep', Nature Biotechnology, 26, 407-415 (2008)

Three compressed downloads are available at this site:

Both demo_limited and demo_full contain a README file that contains instructions how to run miRDeep on your own deep sequencing data. Running it on the demo data allows you to test that it produces what it is supposed to produce (positive control). We highly recommend to test the package on the demo data.

  1. miRDeep.tgz contains the core algorithm script and all auxiliary scripts.
  2. demo_limited.tgz contains all the files that we used to predict C. elegans known and novel miRNA, starting with the file containing the alignments of the filtered reads to the nematode genome and ending with the file of the final predictions (size of this demo: a few MB)
  3. demo_full.tgz contains all the files that we used to predict C. elegans known and novel miRNA, starting with the file containing the alignments of all reads to the nematode genome and ending with the file of the final predictions (warning: size of this file: 100 MB)

For general comments or questions on commercial use, contact: rajewsky at mdc-berlin dot de

(substitute 'at' with '@' and 'dot' with '.' to get the proper email addresses)

Older miRDeep2 versions

pSILAC

 

For the pSILAC webpage follow the link to http://:psilac.mdc-berlin.de