Julia Domingo, Modulation of TF Levels

Precise modulation of transcription factor levels reveals non-linear dosage responses within transcriptional networks

Julia Domingo

She started talking about the genotype-to-phenotype problem. Molecular phenotype vs cellular phenotype. Molecular phenotype – changes in expression of downstream genes. How cis-gene dosage modifies the trans-gene dosage? –> Modulate the expression of few genes and look at the modulation of many targeted gene expressions.

Morris et al, Science, 2023 discovered target genes and pathways at GWAS loci by pooled single-cell CRISPR screens. Some cis-genes are GF1B, NFE2, MYB, TET2.

Excerpts from the talk.

ECCITE-seq framework

HTOs, targeted cDNA, sgRNAs

CRISPRi and CRISPRa

QC and demultiplexing pipeline:

• cDNA
• GDOs (sgRNAs)
• HTOs

Questions and Answers:

1. What is the best unbiased approach to modulate gene expression?

• Wide range of fold changes spanned using different types of sgRNAs. Number of single cells, ON-/OFF- guide target activity don’t affect gene expression.

2. What are the key determinants of dosage effects in cis?

• Distance to TSS, Local chromatin environment

3. How do trans effects relate to dosage changes of the cis gene?

• Cis gene fold change.
• Sigmoid model is a good fit for all the 4 genes (GF1B, NFE2, MYB, TET2).

4. Are transcriptional non-linear dosage response important for downstream phenotype, complex traits or disease?

• Correlation fold changes with mean gene expression. Cell differentiation “pseudophenotype”.
• Disease and GWAS genes are enriched in non-linear responses.

5. Are dosage response of trans genes correlated to gene or network properties?

• Relationship with transcriptional network properties, Mariia Minaeva