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Jason Sheltzer @JSheltzer
, 10 tweets, 4 min read Read on Twitter
Fascinating paper. Knockout mutations that cause mRNA decay trigger the up-regulation of homologous genes, allowing cells to “compensate” for the LOF alteration. People who do CRISPR experiments will have to wrestle with these findings. Some thoughts…

biorxiv.org/content/early/…
I'm worried about multiple-hypothesis testing. That is, they mutate vegfaa, and show that vegfab is regulated. Did they look at vegfb, vegfc, vegfd, etc.? There may be many genes with sequence similarity - how many did they test to find the "compensation" partner?
They show that CRISPR induces compensation, but CRISPRi does not. If this mechanism is generally true, then CRISPRi should be better than CRISPR at finding essential genes and investigating LOF phenotypes. But the opposite has been reported:

nature.com/articles/nbt.3…
Mutations that totally block gene transcription have stronger phenotypes than nonsense mutations that are transcribed. They interpret this as evidence of compensation for the degraded mRNA, but it's also consistent with those mutations being hypomorphs and not nulls.
Their mechanism seems amenable to a big-data approach. We have gene expression and mutation data from hundreds of cancer cell lines - can this pattern be detected? EG are p63 and p73 up-regulated in p53-mutant (but not p53-deleted) cancer cells?
The paper is incredibly cool - injecting uncapped mRNA causes the up-regulation of genes that look like that mRNA! What sort of dark magic-epigenetic-anti-sense-lncRNA fuckery is going on?
So I want to see what happens next! I don't think it's time to throw your KO mouse models out just yet!
Cc @iaincheeseman - very relevant to our conversation earlier this week. Hitting a gene with CRISPR and CRISPRi may cause significantly different phenotypes!
We've generated knockouts of MELK in multiple cancer cell lines. I took a look, and we see no evidence for compensation among MELK's closest homologues (the MARK kinases), among CaMK-family kinases, or among all kinases.
MELK is clearly down-regulated at the RNA level in these clones - evidence of NMD, though we don't know for sure that it's still transcribed at normal levels. Maybe this compensation phenomenon isn't universal? Maybe cancer cells are different than MEFs and zebrafish? TBD!
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More from @JSheltzer see all

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Jim Allison’s immunotherapy Nobel Prize this week underscores the power of a model organism that has attracted a ton of criticism in recent years:
Many papers have highlighted areas where mouse biology significantly differs from human biology. And these are true! Mice are not just furry people.

nytimes.com/2013/02/12/sci…
slate.com/articles/healt…
But mice are mammals that breed rapidly, they can be housed humanely at a pretty low cost, and they have about 97% of the same genes that people do.
Read 13 tweets
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The Nobel Prize in Medicine will be announced this coming Monday. You might think that the winner is a secret, but, with some degree of confidence, you can narrow it down to some likely candidates -
In science, there’s a well-known phenomenon called “the Matthew effect”. In short, the rich get richer. People who successfully get their first grant tend to also get the next one, people who win one prestigious scientific prize tend to win many others. garfield.library.upenn.edu/merton/matthew…
(This phenomenon – and its cousin, the Matilda effect – likely contributes to the lack of female Nobel Laureates. A series of small biases, compounded over time, can lead to immense cumulative disadvantages. journals.sagepub.com/doi/abs/10.117…
Read 13 tweets
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Recently, this remarkable paper went up on @biorxivpreprint – the authors wrote that CRISPR-induced null mutations trigger a pervasive up-regulation of homologs of the deleted gene. It has huge implications for CRISPR technology as well as our understanding of genetic diseases.
My lab makes a lot of null mutations in cancer cells, so it had immediate relevance for our work. I wanted to see if I could observe this phenomenon, so I searched the Gene Expression Omnibus for RNA-Seq experiments using knockout cancer cell lines.
In short, we see no evidence that homologue up-regulation is a common consequence of null mutations in cancer. Here’s an experiment in which ARID1A is knocked out in colon cancer cells, but there’s no up-regulation of its close homologue ARID1B (elifesciences.org/articles/30506).
Read 6 tweets

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Great work as usual by @edyong209 detailing the troubling missteps of the germline #CRISPR editing story of last week. Ed's #3 leaves a lot more to unpack, though, so here's a thread:
theatlantic.com/science/archiv…
Some have interpreted He’s work as a (failed) attempt to replicate the natural 32bp deletion, but I don’t think that was the goal. I think he just hypothesized that any old indel in this genetic region would get the job done and went for it. 2/
Obviously there’s a whole host of things to be concerned about here, but this approach was shockingly cavalier even within the limited scope of what’s known about loss of CCR5 function. 3/
Read 16 tweets
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So let's comment on these reports that came out today on the creation of the first #CRISPR babies from a team at Shenzhen, China reported here -> technologyreview.com/s/612458/exclu… or here -> apnews.com/4997bb7aa36c45… and this Youtube video
What we know about it? Well not much. 1/ They have edited CCR5 gene, which prevent HIV entry. Have they edited CCR5 to generate a ∆32 mutation? no idea. It looks they edited CCR5 in the pronuclear and check editing by sequencing 3-5 days later -> 16/22 edited and 11 implanted
Twins were born with one carrying an heterozygous allele and the second baby an homozygous allele and one carried mosaic alleles. Details? No off target found but I don't have any details on this too. Are the CCR5 protein level changed? Again very little details on this.
Read 10 tweets
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Finally the comments to Mitalipov’s group paper on #CRISPR in human embryos are out in @nature as well as the rebuttal from Ma et al. as brief communications arising #paywall. Let’s have a look more in details -> Thread
One of the major finding of the Ma et al. paper was the discovery of high frequency of inter-homologous repair mechanisms aka loss of heterozygocity LOH -> 1 allele is used as a template for
#CRISPR-HDR -> WT alleles are in fact alleles from the maternal alleles only.
We have been debating a lot on this in the last year or so. The first important comment from @Fatwa_Adikusuma et al is Ma et al. missed deleted alleles. In the paper they edited 6 loci in zygotes and EScells and checked for large deletions nature.com/articles/s4158…
Read 15 tweets
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Daniel Weinberger: been arguing for years that to understand schizophrenia we need to think about how brains are built and how different trajectories of post natal development affect the brain #BAP2018
Weinberger: schizophrenia not a disease in a classical medical sense but a state of brain formed through development #BAP2018
Weinberger: Schizophrenia runs in families. Roughly 1% of cases have loss of function mutations which is major risk factor, about 2% cases have CNVs and 97% cases cumulation of common variants in particular combination. #BAP2018
Read 19 tweets
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Hace una semana lanzé un HILO en el que explicaba los orígenes e información básica para entender las herramientas #CRISPR. Gracias a @jm_alarcon ese HILO está recogido en esta web: threadreaderapp.com/thread/9834491… Hoy continuaré con otro en el que hablaré de las aplicaciones. Dentro HILO
Las herramientas #CRISPR nos han cambiado tanto la vida a los investigadores de ciencias de la vida que prácticamente suelo decir que pronto solamente habrá dos tipos de laboratorio: (1) los que YA están usando CRISPR en sus experimentos y (2) los que VAN a usar CRISPR. Y ya está
Lo que está ocurriendo con las #CRISPR y la edición genética de alguna manera recuerda a lo que ocurrió a finales de los años 80, cuando apareció y se instaló la técnica de amplificación del ADN #PCR (reacción de la polimerasa en cadena) que está presente hoy en todas partes.
Read 38 tweets
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En apenas 5 años (los primeros experimentos que demostraban su uso como editores genéticos aparecieron en enero de 2013) las herramientas #CRISPR han revolucionado los laboratorios de todo el mundo. Voy a intentar explicar por qué, de forma resumida. Vamos con el prometido HILO:
Voy a empezar por el final. Lo que hace a las #CRISPR unas herramientas sorprendentes es su aparente facilidad para promover cambios en el genoma, en el material genético de cualquier organismo. Recordemos que el ADN está formado por cadenas de letras (nucleótidos) A, T, G y C
El ADN es una doble cadena. La A siempre se empareja con la T, y la G con la C. Cada organismo tiene un número de letras específico. Nosotros tenemos aproximadamente 3.000 millones de letras. Por el contrario una bacteria como Escherichia coli tiene apenas 3 millones de letras.
Read 38 tweets

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