Spatial reconstruction of single enterocytes uncovers broad zonation along the intestinal villus axis

Abstract

The intestinal epithelium is a highly structured tissue composed of repeating crypt-villus units1,2. Enterocytes, which constitute the most abundant cell type, perform the diverse tasks of absorbing a wide range of nutrients while protecting the body from the harsh bacterial-rich environment. It is unknown if these tasks are equally performed by all enterocytes or whether they are spatially zonated along the villus axis3. Here, we performed whole-transcriptome measurements of laser-capture-microdissected villus segments to extract a large panel of landmark genes, expressed in a zonated manner. We used these genes to localize single sequenced enterocytes along the villus axis, thus reconstructing a global spatial expression map. We found that most enterocyte genes were zonated. Enterocytes at villi bottoms expressed an anti-bacterial Reg gene program in a microbiome-dependent manner, potentially reducing the crypt pathogen exposure. Translation, splicing and respiration genes steadily decreased in expression towards the villi tops, whereas distinct mid-top villus zones sub-specialized in the absorption of carbohydrates, peptides and fat. Enterocytes at the villi tips exhibited a unique gene-expression signature consisting of Klf4, Egfr, Neat1, Malat1, cell adhesion and purine metabolism genes. Our study exposes broad spatial heterogeneity of enterocytes, which could be important for achieving their diverse tasks.

Stem cell lineage survival as a noisy competition for niche access

“…For example, the prediction of the number of stem cells in the crypt, even though accurate with respect to live-imaging functional data, does not explain why Lgr5 levels display a clear on/off pattern inside vs. outside the crypt, and reversal to a stem cell fate is not always a fast process depen-dent solely on niche cues. Therefore, a natural follow up should explore the relation between the emergent properties of the dynamics and the molecular markers, jointly to their functional differentiation [43,44]. In addition, more complex geometries, and with that, other tissues, could be explored within our framework.…”

Spatial sorting enables comprehensive characterization of liver zonation

“…We demonstrated applications of this approach for resolving the zonation of hepatocyte proteins and miRs. The approach can be readily applied to other structured organs and cells types exhibiting zonation, including liver endothelial cells 11 , intestinal enterocytes 46 and kidney cells 47,48 . The usage of endogenous surface markers renders spatial sorting particularly useful for studying zonation in humans as well.…”

CellTag Indexing: genetic barcode-based sample multiplexing for single-cell genomics

“…Thus, to capture the full spectrum of intestinal cell identities, we opted to use whole tissue single-nucleus extraction, over epithelial isolation and digestion, to process the engrafted colon for RNA-sequencing. Indeed, single- [25][26][27][28] , Reg4 in deep crypt secretory cells 29 , Myt1l, Asic2, and Syt1 in enteric neurons [30][31][32][33] , Vil1, Plac8, and Krt20 in enterocytes [34][35][36] , Nkx2-2, Chga, and Tph1 in enteroendocrine cells 37,38 , and Fcgbp, Muc2, and Clca1 in goblet cells 39,40 (Figs. 2G and S5C-E).…”