Machine-learning-optimized Cas12a barcoding enables the recovery of single-cell lineages and transcriptional profiles

Hughes, Nicholas W.; Qu, Yuanhao; Zhang, Jiaqi; Tang, Weijing; Pierce, Justin; Wang, Chengkun; Agrawal, Aditi; Morri, Maurizio; Neff, Norma; Winslow, Monte M.; Wang, Mengdi; Cong, Le

doi:10.1016/j.molcel.2022.06.001

Cited by 20 publications

(20 citation statements)

References 75 publications

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“…which is hereinafter referred to as the "memory index". We note that this definition of memory index is similar to that used in previous transcriptional memory-related studies 29,39 .…”

Section: Transcriptional Memory Has Limited Contribution To Developme...mentioning

confidence: 88%

“…It is imperative to note that as our hypothesis revolves around the above-mentioned functional units, CLTs with sufficient resolution (fraction of cells sampled) are essential, otherwise stereotyped development cannot be detected with only <1% cells sampled from each functional unit. In addition, a high resolution CLT would also reveal how stereotyped development occurs, such as mitotic-coupling versus population-coupling development 18 and whether epigenetic memory 29 plays a role.…”

Section: Introductionmentioning

confidence: 99%

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Stereotyped cell lineage trees support robust development

Zhang,

Li,

Chen

et al. 2024

Preprint

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Multicellular organisms must have robust development to ensure physiological stability in the face of environmental changes or perturbations. While various mechanisms contributing to developmental robustness have been identified at the subcellular level, those at the intercellular and tissue level remain largely unknown. Our study explores this question using an in vitro directed differentiation model of human embryonic stem cells (hESCs) into lung progenitor cells. Integrated analysis of single-cell transcriptomes and high-density cell lineage trees (CLTs) of the same colonies allowed a fine-resolution recapitulation of known cell types, as well as their differentiation hierarchies and developmental trajectories. Most importantly, we observed stable cell type compositions among many sub-CLTs across biological replicates. Systematic comparison among CLTs by a novel computational framework for CLT alignment suggests that stereotypical development extends beyond stable cell type composition to a degree of significant resemblance in sub-CLT topology. The existence of such sub-CLTs resembling each other not only deepens our understanding of developmental robustness by demonstrating the existence of a stereotyped program, but also suggests a novel perspective for the function of specific cell types in the context of stereotyped sub-CLTs.

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“…which is hereinafter referred to as the "memory index". We note that this definition of memory index is similar to that used in previous transcriptional memory-related studies 29,39 .…”

Section: Transcriptional Memory Has Limited Contribution To Developme...mentioning

confidence: 88%

Section: Introductionmentioning

confidence: 99%

Stereotyped cell lineage trees support robust development

Zhang,

Li,

Chen

et al. 2024

Preprint

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show abstract

“…Despite the rapid development of CRISPR lineage tracing methods (4,29,60), building highprecision lineage trees with single-cell resolution is still challenging because of the small number of Cas9 target sites (typically<50), rapid saturation, frequent inter-site deletions, and other factors (61)(62)(63). This requires a more reliable lineage tracing method that has a larger lineage-labeling space and more stable mutagenesis strategy.…”

Section: Discussionmentioning

confidence: 99%

Cell division history encodes directional information of fate transitions

Wang

Hou

Lü

et al. 2022

Preprint

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Single-cell RNA-sequencing (scRNA-seq) enables systematic mapping of cellular differentiation trajectories. However, inferring the cell-fate transitions under diseases or perturbations is still challenging due to the high cellular plasticity. Here, we demonstrate that monotonically expressed genes (MEGs) along cell divisions record the directions of state transitions regardless of the cellular processes. We developed a computational framework (PhyloVelo) to identify MEGs and reconstruct a novel transcriptomic velocity field by leveraging both scRNA-seq and phylogenetic information. PhyloVelo accurately recovered linear, bifurcated and convergent differentiations in simulations and C. elegans. It outperformed current approaches for delineating cellular trajectories in embryo development and tumor evolution through analysis of five CRISPR/Cas9-based lineage tracing datasets. Together, our study unveils an internal cellular clock and provides a powerful method for cell-fate analysis in diverse biological contexts.

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“…This is ameliorated using a system like Tickertape/DNA Typewriter [101, 102] whereby edits in an array happen sequentially, instead of randomly as in GESTALT. DAISY uses an alternate approach employing Cas12a and dual acting inverted site arrays that enable higher entropy and ∼66,000 potential barcodes in a smaller transgene package [103]. It should be noted that not many of these systems have been examined with spatial transcriptomics.…”

Section: Expanding Beyond Genetic Reporter Genes With Large-scale Gen...mentioning

confidence: 99%

Beyond the Rainbow: A Review of Advanced Lineage Tracing Methodologies for Interrogating the Initiation, Evolution, and Recurrence of Brain Tumors

Sabet¹,

Breunig²

2023

Dev Neurosci

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The mammalian forebrain is perhaps the pinnacle of evolution and one of the most complex structures in known existence. The origin of this complexity and diversity partly lies in dynamic behavior of progenitors during embryonic neural development, all of which is under the control of regulatory mechanisms that ensure all the elements end up in the right place at the right time. Historically, dye-base, histochemical, enzymatic, or fluorescent lineage tracing techniques have been used deconvolute developmental dynamics in tissues and cells. Technical limitations resulted from a restrictive number of fluorophores, the half-life of the dyes, or the ability to deconvolute mixed population. These limitations often impede larger scale lineage tracing using these methods in spatial and temporal contexts. Genetic barcoding techniques have been used for decades to explore clonal investigations and have now evolved with high-throughput sequencing methods to allow for impressive insights into population and even organism level lineage relationships. In this review we will discuss the contemporary progression of lineage tracing methodologies and how they were applied to answer questions around molecular and cellular mechanisms of gliogenesis and neurogenesis. We will also discuss recent advances in computational biology, single-cell sequencing, and in situ-based lineage tracing methodologies. Incorporation of these methods into toolset of lineage tracing promise to enable a higher-resolution, multimodal view of neural lineages during development and disease processes that highjack developmental signaling such as brain tumor development and recurrence—where traditional developmental hierarchies become more plastic and less predictable. Given the dismal prognosis of high-grade brain tumors like glioblastoma multiforme, a better understanding of the lineage relationships leading to disease heterogeneity and recurrence is desperately needed to formulate efficacious approaches to treatment. Here we discuss the past, present, and future of lineage tracing at the intersection of development and disease.

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Machine-learning-optimized Cas12a barcoding enables the recovery of single-cell lineages and transcriptional profiles

Cited by 20 publications

References 75 publications

Stereotyped cell lineage trees support robust development

Stereotyped cell lineage trees support robust development

Cell division history encodes directional information of fate transitions

Beyond the Rainbow: A Review of Advanced Lineage Tracing Methodologies for Interrogating the Initiation, Evolution, and Recurrence of Brain Tumors

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