Close

Long-term live imaging of the Drosophila adult midgut reveals real-time dynamics of cell division, differentiation, and loss

Judy Martin, Erin Nicole Sanders, Paola Moreno-Roman, Shruthi Balachandra, XinXin Du, Leslie Ann Jaramillo Koyama, Lucy Erin O'Brien

Preprint posted on 26 February 2018 https://www.biorxiv.org/content/early/2018/02/26/271742

Article now published in eLife at http://dx.doi.org/10.7554/elife.36248

A new method captures the dynamics of tissue homeostasis in the stem-cell based organ of the adult fly midgut

Selected by Natalie Dye

Background:

To maintain tissue homeostasis, many adult organs harbour pools of stem cells that divide and differentiate to replenish cells that are lost through ageing and/or injury. How these division and differentiation events are regulated is an important open question in biology.

Due to its genetic tractability, the adult Drosophila midgut (analogous to the mammalian small intestine) has been a powerful model system for identifying the conserved signalling pathways underlying both normal tissue homeostasis and its dysfunction during disease, infection, and ageing. Nonetheless, without methods to support extended live imaging of this tissue (longer than 90min), it has been difficult to use this powerful genetic model to study the cell biology and signalling dynamics underlying stem-cell-based homeostasis.

Summary of the preprint:

The authors of this preprint create a new and vastly improved protocol for imaging the midgut. They first stabilize the abdomen of the animal while providing a feeding tube; then they remove part of the dorsal cuticle, immerse this exposed part in media, and image through this media (Fig 1). The authors develop an image processing pipeline to compensate for global movements of the tissue and use a combination of fluorescent genetic markers and quantification of nuclear size to simultaneously identify and quantify the dynamics of different midgut cell types on a tissue scale for 12-16 hrs.

Fig 1: Method schematic (Fig 1A-C of the preprint): (A) Illustration of the fly digestive track; midgut in white. (B) Mounting the animal for imaging (with an upright microscope). The midgut is accessed through a hole in the cuticle, and the animal is fed through a tube. (C) Steps in sample preparation.

Movie S5 from the preprint showing the a ‘fate sensor’ midgut (esgGal4, UAS-his2b::CFP, GBE-Su(H)-GFP:nls; ubi-his2av::mRFP). With the combination of nuclear size and genetic labels, four midgut cell types can be identified and tracked: stem cells (red), enteroblasts (yellow-green), enterocytes (gray, polyploid – and thus larger), and enteroendocrine cells (gray, diploid – and thus smaller).

 

To demonstrate the power of their technique, the authors then provide new observations related to three processes occurring during normal tissue homeostasis:

  • Loss. Novel observations regarding the extrusion of enterocytes (terminally differentiated, nutrient-absorbing cells) include:
    1. Basal cross sectional cell area shrinks in a rachet-like pulsatile fashion
    2. The nucleus rapidly shoots out of the tissue toward the lumen before recoiling back toward the epithelial surface.
  • Division. Stem cell divisions occur less frequently during live imaging than would be expected based on staining of fixed tissues. Nonetheless, the authors report observations from 39 mitosis events—far more than ever before. Results not only confirm previous work using fixed tissues but also include the following new observations:
    1. Dividing stem cells can rapidly reorient before division.
    2. Neighbouring enteroblasts influence stem cell division orientation.
  • Differentiation. While genetic experiments have clearly implicated Notch signalling in stem cell differentiation, without live imaging data, the dynamics have been elusive, limiting our understanding of the regulatory mechanisms. Here, the authors calibrate a live Notch activity reporter and make three new discoveries:
    1. Notch activation and stem cell differentiation can be directly observed live (Fig 2) and occurs over a time-scale of hours (based on observation of 5 events).
    2. Proximity between stem cell siblings after division does not correlate with Notch activity.
    3. The kinetics of Notch activation and cell differentiation imply a latency period after a cell is born, whereby its differentiation is delayed for several hours.

Fig 2: Direct observation of Notch activation during cell fate transitions (Fig 5D from the preprint). GBE-Su(H)-GFP:nls reports Notch transcriptional activity, while ubi-his2av::mRFP provides a reference signal. Cells additionally express esg>his2ab::CFP to mark progenitor cells. The authors use the GFP:RFP ratio to quantify Notch activation and directly observe transitions between stem and enteroblast states, finding that these transitions require several hours (2.4-6.9hrs).

 

Why I chose this preprint:

In my humble opinion, methods papers are where the preprint servers are especially valuable. Getting a new method out in the open as soon as possible is a great thing. Included in this paper are videos and detailed diagrams to help the reader use the method. Slight modifications may enable imaging of different parts of the adult fly, making the approach potentially even more widely applicable. Drosophila genetics has proven to be extremely powerful over the decades, but I’m excited to see more and more recent efforts to add live imaging to the toolbox – from the embryo, to the brain, wing disc, and now gut.

While this paper is first and foremost a method paper, there’s also a lot of tantalizing new biology presented. Their new data demonstrate how certain kinds of information can really only be obtained by direct observation over time, something I myself have been aiming for in my own work. The authors provide solid, valuable characterization of the dynamics—but nevertheless their data seem to generate more questions than answers. Thus, I think the best is yet to come. There is great potential for the live imaging approach to synergize with existing genetic tools in Drosophila to make really fundamental mechanistic discoveries. In principle, their techniques may also be combined with laser ablation, to probe the influence of mechanical forces, and pharmacological perturbations to rapidly, locally, and reversibly inhibit various proteins.

 

Future Directions:

Here are some of the questions I’m looking forward to hearing about in the future:

  • What in the world is going on with the nucleus in those extruding cells?
  • Since the duration of stem cell sibling contact (at least at the level of the nucleus) does not seem to be important, what regulates the when and whether Notch is activated?
    • Related question: are there filopodia-like protrusions in these cells that may signal long distances through Notch, as in the Drosophila notum?
  • What is the molecular mechanism underlying the apparent latency phase in stem cell differentiation?
  • What happens to these dynamics upon injury, infection, ageing, and with varying diets?
    • Related comment: another recent preprint demonstrates that dietary lipids affect cell differentiation and Notch signalling in the midgut (Obniski et al 2018: https://www.biorxiv.org/content/early/2018/02/28/273813). This is just one example of the interesting perturbations that can be now studied live using the techniques from O’Brien and colleagues.

 

Further reading:

  • Cohen M, Georgiou M, Stevenson NL, Miodownik M, Baum B. “Dynamic filopodia transmit intermittent Delta-Notch signaling to drive pattern refinement during lateral inhibition.” Dev Cell. 2010. doi: 10.1016/j.devcel.2010.06.006.
  • Gervais L, Bardin AJ. “Tissue homeostasis and aging: new insight from the fly intestine.” Curr Opin Cell Biol. 2017. doi: 10.1016/j.ceb.2017.06.005.
  • Guo Z, Lucchetta E, Rafel N, Ohlstein B.“Maintenance of the adult Drosophila intestine: all roads lead to homeostasis.” Curr Opin Genet Dev. 2016. doi: 10.1016/j.gde.2016.06.009.
  • Obniski R, Sieber M, and Spradling A. “Dietary lipids modulate Notch signaling and influence adult intestinal development and metabolism in Drosophila.” BioRxiv 2018. https://www.biorxiv.org/content/early/2018/02/28/273813

 

Tags: cell extrusion, differentiation, homeostasis, intestine, microscopy, notch, organ renewal, stem cells

Posted on: 20 March 2018

Read preprint (No Ratings Yet)

Have your say

Your email address will not be published. Required fields are marked *

This site uses Akismet to reduce spam. Learn how your comment data is processed.

Sign up to customise the site to your preferences and to receive alerts

Register here

preLists in the cancer biology category:

CSHL 87th Symposium: Stem Cells

Preprints mentioned by speakers at the #CSHLsymp23

 



List by Alex Eve

Journal of Cell Science meeting ‘Imaging Cell Dynamics’

This preList highlights the preprints discussed at the JCS meeting 'Imaging Cell Dynamics'. The meeting was held from 14 - 17 May 2023 in Lisbon, Portugal and was organised by Erika Holzbaur, Jennifer Lippincott-Schwartz, Rob Parton and Michael Way.

 



List by Helen Zenner

CellBio 2022 – An ASCB/EMBO Meeting

This preLists features preprints that were discussed and presented during the CellBio 2022 meeting in Washington, DC in December 2022.

 



List by Nadja Hümpfer et al.

Fibroblasts

The advances in fibroblast biology preList explores the recent discoveries and preprints of the fibroblast world. Get ready to immerse yourself with this list created for fibroblasts aficionados and lovers, and beyond. Here, my goal is to include preprints of fibroblast biology, heterogeneity, fate, extracellular matrix, behavior, topography, single-cell atlases, spatial transcriptomics, and their matrix!

 



List by Osvaldo Contreras

Single Cell Biology 2020

A list of preprints mentioned at the Wellcome Genome Campus Single Cell Biology 2020 meeting.

 



List by Alex Eve

ASCB EMBO Annual Meeting 2019

A collection of preprints presented at the 2019 ASCB EMBO Meeting in Washington, DC (December 7-11)

 



List by Madhuja Samaddar et al.

Lung Disease and Regeneration

This preprint list compiles highlights from the field of lung biology.

 



List by Rob Hynds

Anticancer agents: Discovery and clinical use

Preprints that describe the discovery of anticancer agents and their clinical use. Includes both small molecules and macromolecules like biologics.

 



List by Zhang-He Goh

Biophysical Society Annual Meeting 2019

Few of the preprints that were discussed in the recent BPS annual meeting at Baltimore, USA

 



List by Joseph Jose Thottacherry

Also in the cell biology category:

‘In preprints’ from Development 2022-2023

A list of the preprints featured in Development's 'In preprints' articles between 2022-2023

 



List by Alex Eve, Katherine Brown

preLights peer support – preprints of interest

This is a preprint repository to organise the preprints and preLights covered through the 'preLights peer support' initiative.

 



List by preLights peer support

The Society for Developmental Biology 82nd Annual Meeting

This preList is made up of the preprints discussed during the Society for Developmental Biology 82nd Annual Meeting that took place in Chicago in July 2023.

 



List by Joyce Yu, Katherine Brown

CSHL 87th Symposium: Stem Cells

Preprints mentioned by speakers at the #CSHLsymp23

 



List by Alex Eve

Journal of Cell Science meeting ‘Imaging Cell Dynamics’

This preList highlights the preprints discussed at the JCS meeting 'Imaging Cell Dynamics'. The meeting was held from 14 - 17 May 2023 in Lisbon, Portugal and was organised by Erika Holzbaur, Jennifer Lippincott-Schwartz, Rob Parton and Michael Way.

 



List by Helen Zenner

9th International Symposium on the Biology of Vertebrate Sex Determination

This preList contains preprints discussed during the 9th International Symposium on the Biology of Vertebrate Sex Determination. This conference was held in Kona, Hawaii from April 17th to 21st 2023.

 



List by Martin Estermann

Alumni picks – preLights 5th Birthday

This preList contains preprints that were picked and highlighted by preLights Alumni - an initiative that was set up to mark preLights 5th birthday. More entries will follow throughout February and March 2023.

 



List by Sergio Menchero et al.

CellBio 2022 – An ASCB/EMBO Meeting

This preLists features preprints that were discussed and presented during the CellBio 2022 meeting in Washington, DC in December 2022.

 



List by Nadja Hümpfer et al.

Fibroblasts

The advances in fibroblast biology preList explores the recent discoveries and preprints of the fibroblast world. Get ready to immerse yourself with this list created for fibroblasts aficionados and lovers, and beyond. Here, my goal is to include preprints of fibroblast biology, heterogeneity, fate, extracellular matrix, behavior, topography, single-cell atlases, spatial transcriptomics, and their matrix!

 



List by Osvaldo Contreras

EMBL Synthetic Morphogenesis: From Gene Circuits to Tissue Architecture (2021)

A list of preprints mentioned at the #EESmorphoG virtual meeting in 2021.

 



List by Alex Eve

FENS 2020

A collection of preprints presented during the virtual meeting of the Federation of European Neuroscience Societies (FENS) in 2020

 



List by Ana Dorrego-Rivas

Planar Cell Polarity – PCP

This preList contains preprints about the latest findings on Planar Cell Polarity (PCP) in various model organisms at the molecular, cellular and tissue levels.

 



List by Ana Dorrego-Rivas

BioMalPar XVI: Biology and Pathology of the Malaria Parasite

[under construction] Preprints presented at the (fully virtual) EMBL BioMalPar XVI, 17-18 May 2020 #emblmalaria

 



List by Dey Lab, Samantha Seah

1

Cell Polarity

Recent research from the field of cell polarity is summarized in this list of preprints. It comprises of studies focusing on various forms of cell polarity ranging from epithelial polarity, planar cell polarity to front-to-rear polarity.

 



List by Yamini Ravichandran

TAGC 2020

Preprints recently presented at the virtual Allied Genetics Conference, April 22-26, 2020. #TAGC20

 



List by Maiko Kitaoka et al.

3D Gastruloids

A curated list of preprints related to Gastruloids (in vitro models of early development obtained by 3D aggregation of embryonic cells). Updated until July 2021.

 



List by Paul Gerald L. Sanchez and Stefano Vianello

ECFG15 – Fungal biology

Preprints presented at 15th European Conference on Fungal Genetics 17-20 February 2020 Rome

 



List by Hiral Shah

ASCB EMBO Annual Meeting 2019

A collection of preprints presented at the 2019 ASCB EMBO Meeting in Washington, DC (December 7-11)

 



List by Madhuja Samaddar et al.

EMBL Seeing is Believing – Imaging the Molecular Processes of Life

Preprints discussed at the 2019 edition of Seeing is Believing, at EMBL Heidelberg from the 9th-12th October 2019

 



List by Dey Lab

Autophagy

Preprints on autophagy and lysosomal degradation and its role in neurodegeneration and disease. Includes molecular mechanisms, upstream signalling and regulation as well as studies on pharmaceutical interventions to upregulate the process.

 



List by Sandra Malmgren Hill

Lung Disease and Regeneration

This preprint list compiles highlights from the field of lung biology.

 



List by Rob Hynds

Cellular metabolism

A curated list of preprints related to cellular metabolism at Biorxiv by Pablo Ranea Robles from the Prelights community. Special interest on lipid metabolism, peroxisomes and mitochondria.

 



List by Pablo Ranea Robles

BSCB/BSDB Annual Meeting 2019

Preprints presented at the BSCB/BSDB Annual Meeting 2019

 



List by Dey Lab

MitoList

This list of preprints is focused on work expanding our knowledge on mitochondria in any organism, tissue or cell type, from the normal biology to the pathology.

 



List by Sandra Franco Iborra

ASCB/EMBO Annual Meeting 2018

This list relates to preprints that were discussed at the recent ASCB conference.

 



List by Dey Lab, Amanda Haage

Also in the developmental biology category:

GfE/ DSDB meeting 2024

This preList highlights the preprints discussed at the 2024 joint German and Dutch developmental biology societies meeting that took place in March 2024 in Osnabrück, Germany.

 



List by Joyce Yu

‘In preprints’ from Development 2022-2023

A list of the preprints featured in Development's 'In preprints' articles between 2022-2023

 



List by Alex Eve, Katherine Brown

preLights peer support – preprints of interest

This is a preprint repository to organise the preprints and preLights covered through the 'preLights peer support' initiative.

 



List by preLights peer support

The Society for Developmental Biology 82nd Annual Meeting

This preList is made up of the preprints discussed during the Society for Developmental Biology 82nd Annual Meeting that took place in Chicago in July 2023.

 



List by Joyce Yu, Katherine Brown

CSHL 87th Symposium: Stem Cells

Preprints mentioned by speakers at the #CSHLsymp23

 



List by Alex Eve

Journal of Cell Science meeting ‘Imaging Cell Dynamics’

This preList highlights the preprints discussed at the JCS meeting 'Imaging Cell Dynamics'. The meeting was held from 14 - 17 May 2023 in Lisbon, Portugal and was organised by Erika Holzbaur, Jennifer Lippincott-Schwartz, Rob Parton and Michael Way.

 



List by Helen Zenner

9th International Symposium on the Biology of Vertebrate Sex Determination

This preList contains preprints discussed during the 9th International Symposium on the Biology of Vertebrate Sex Determination. This conference was held in Kona, Hawaii from April 17th to 21st 2023.

 



List by Martin Estermann

Alumni picks – preLights 5th Birthday

This preList contains preprints that were picked and highlighted by preLights Alumni - an initiative that was set up to mark preLights 5th birthday. More entries will follow throughout February and March 2023.

 



List by Sergio Menchero et al.

CellBio 2022 – An ASCB/EMBO Meeting

This preLists features preprints that were discussed and presented during the CellBio 2022 meeting in Washington, DC in December 2022.

 



List by Nadja Hümpfer et al.

2nd Conference of the Visegrád Group Society for Developmental Biology

Preprints from the 2nd Conference of the Visegrád Group Society for Developmental Biology (2-5 September, 2021, Szeged, Hungary)

 



List by Nándor Lipták

Fibroblasts

The advances in fibroblast biology preList explores the recent discoveries and preprints of the fibroblast world. Get ready to immerse yourself with this list created for fibroblasts aficionados and lovers, and beyond. Here, my goal is to include preprints of fibroblast biology, heterogeneity, fate, extracellular matrix, behavior, topography, single-cell atlases, spatial transcriptomics, and their matrix!

 



List by Osvaldo Contreras

EMBL Synthetic Morphogenesis: From Gene Circuits to Tissue Architecture (2021)

A list of preprints mentioned at the #EESmorphoG virtual meeting in 2021.

 



List by Alex Eve

EMBL Conference: From functional genomics to systems biology

Preprints presented at the virtual EMBL conference "from functional genomics and systems biology", 16-19 November 2020

 



List by Jesus Victorino

Single Cell Biology 2020

A list of preprints mentioned at the Wellcome Genome Campus Single Cell Biology 2020 meeting.

 



List by Alex Eve

Society for Developmental Biology 79th Annual Meeting

Preprints at SDB 2020

 



List by Irepan Salvador-Martinez, Martin Estermann

FENS 2020

A collection of preprints presented during the virtual meeting of the Federation of European Neuroscience Societies (FENS) in 2020

 



List by Ana Dorrego-Rivas

Planar Cell Polarity – PCP

This preList contains preprints about the latest findings on Planar Cell Polarity (PCP) in various model organisms at the molecular, cellular and tissue levels.

 



List by Ana Dorrego-Rivas

Cell Polarity

Recent research from the field of cell polarity is summarized in this list of preprints. It comprises of studies focusing on various forms of cell polarity ranging from epithelial polarity, planar cell polarity to front-to-rear polarity.

 



List by Yamini Ravichandran

TAGC 2020

Preprints recently presented at the virtual Allied Genetics Conference, April 22-26, 2020. #TAGC20

 



List by Maiko Kitaoka et al.

3D Gastruloids

A curated list of preprints related to Gastruloids (in vitro models of early development obtained by 3D aggregation of embryonic cells). Updated until July 2021.

 



List by Paul Gerald L. Sanchez and Stefano Vianello

ASCB EMBO Annual Meeting 2019

A collection of preprints presented at the 2019 ASCB EMBO Meeting in Washington, DC (December 7-11)

 



List by Madhuja Samaddar et al.

EDBC Alicante 2019

Preprints presented at the European Developmental Biology Congress (EDBC) in Alicante, October 23-26 2019.

 



List by Sergio Menchero et al.

EMBL Seeing is Believing – Imaging the Molecular Processes of Life

Preprints discussed at the 2019 edition of Seeing is Believing, at EMBL Heidelberg from the 9th-12th October 2019

 



List by Dey Lab

SDB 78th Annual Meeting 2019

A curation of the preprints presented at the SDB meeting in Boston, July 26-30 2019. The preList will be updated throughout the duration of the meeting.

 



List by Alex Eve

Lung Disease and Regeneration

This preprint list compiles highlights from the field of lung biology.

 



List by Rob Hynds

Young Embryologist Network Conference 2019

Preprints presented at the Young Embryologist Network 2019 conference, 13 May, The Francis Crick Institute, London

 



List by Alex Eve

Pattern formation during development

The aim of this preList is to integrate results about the mechanisms that govern patterning during development, from genes implicated in the processes to theoritical models of pattern formation in nature.

 



List by Alexa Sadier

BSCB/BSDB Annual Meeting 2019

Preprints presented at the BSCB/BSDB Annual Meeting 2019

 



List by Dey Lab

Zebrafish immunology

A compilation of cutting-edge research that uses the zebrafish as a model system to elucidate novel immunological mechanisms in health and disease.

 



List by Shikha Nayar

Also in the genetics category:

9th International Symposium on the Biology of Vertebrate Sex Determination

This preList contains preprints discussed during the 9th International Symposium on the Biology of Vertebrate Sex Determination. This conference was held in Kona, Hawaii from April 17th to 21st 2023.

 



List by Martin Estermann

Alumni picks – preLights 5th Birthday

This preList contains preprints that were picked and highlighted by preLights Alumni - an initiative that was set up to mark preLights 5th birthday. More entries will follow throughout February and March 2023.

 



List by Sergio Menchero et al.

Semmelweis Symposium 2022: 40th anniversary of international medical education at Semmelweis University

This preList contains preprints discussed during the 'Semmelweis Symposium 2022' (7-9 November), organised around the 40th anniversary of international medical education at Semmelweis University covering a wide range of topics.

 



List by Nándor Lipták

20th “Genetics Workshops in Hungary”, Szeged (25th, September)

In this annual conference, Hungarian geneticists, biochemists and biotechnologists presented their works. Link: http://group.szbk.u-szeged.hu/minikonf/archive/prg2021.pdf

 



List by Nándor Lipták

2nd Conference of the Visegrád Group Society for Developmental Biology

Preprints from the 2nd Conference of the Visegrád Group Society for Developmental Biology (2-5 September, 2021, Szeged, Hungary)

 



List by Nándor Lipták

EMBL Conference: From functional genomics to systems biology

Preprints presented at the virtual EMBL conference "from functional genomics and systems biology", 16-19 November 2020

 



List by Jesus Victorino

TAGC 2020

Preprints recently presented at the virtual Allied Genetics Conference, April 22-26, 2020. #TAGC20

 



List by Maiko Kitaoka et al.

ECFG15 – Fungal biology

Preprints presented at 15th European Conference on Fungal Genetics 17-20 February 2020 Rome

 



List by Hiral Shah

Autophagy

Preprints on autophagy and lysosomal degradation and its role in neurodegeneration and disease. Includes molecular mechanisms, upstream signalling and regulation as well as studies on pharmaceutical interventions to upregulate the process.

 



List by Sandra Malmgren Hill
Close