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A library of mammalian effector modules for synthetic morphology.

Cachat E, Liu W, Hohenstein P, Davies JA - J Biol Eng (2014)

Bottom Line: Together with cell differentiation, these mechanisms allow populations of cells to organize themselves into defined geometries and structures, as simple embryos develop into complex organisms.Here we describe this library and demonstrate its use in the T-REx-293 human cell line to induce each of these desired morphological behaviours on command.Building on from the simple test systems described here, we want to extend engineered control of morphogenetic cell behaviour to more complex 3D structures that can inform embryologists and may, in the future, be used in surgery and regenerative medicine, making synthetic morphology a powerful tool for developmental biology and tissue engineering.

View Article: PubMed Central - PubMed

Affiliation: University of Edinburgh, Centre for Integrative Physiology, Hugh Robson Building, George Square, Edinburgh, EH8 9XD UK.

ABSTRACT

Background: In mammalian development, the formation of most tissues is achieved by a relatively small repertoire of basic morphogenetic events (e.g. cell adhesion, locomotion, apoptosis, etc.), permutated in various sequences to form different tissues. Together with cell differentiation, these mechanisms allow populations of cells to organize themselves into defined geometries and structures, as simple embryos develop into complex organisms. The control of tissue morphogenesis by populations of engineered cells is a potentially very powerful but neglected aspect of synthetic biology.

Results: We have assembled a modular library of synthetic morphogenetic driver genes to control (separately) mammalian cell adhesion, locomotion, fusion, proliferation and elective cell death. Here we describe this library and demonstrate its use in the T-REx-293 human cell line to induce each of these desired morphological behaviours on command.

Conclusions: Building on from the simple test systems described here, we want to extend engineered control of morphogenetic cell behaviour to more complex 3D structures that can inform embryologists and may, in the future, be used in surgery and regenerative medicine, making synthetic morphology a powerful tool for developmental biology and tissue engineering.

No MeSH data available.


Related in: MedlinePlus

Enhanced locomotion induced by tetracycline in T-REx-293 cells engineered withCrk-II.(a) DAPI staining (blue) and phalloidin-FITC (green) during scratch closing assays showing enhanced lamellipodia formation in THLOII-15 cells (a representative clone of T-REx-293 cells carrying the locomotion module) when induced with tetracycline: the induced cells show more frequent formation of lamellipodia, shown in green. Scale bars: 100 μm. (b) Distance travelled by cells in scratch closing assays with or without induction of the locomotion module. Images were acquired every 10 min and coordinates of 4 groups of cells on each side of wounds were recorded (6 to 18 h post-induction). Tetracycline increased the migration of THLOII-15 cells (trail plot and graph, *p <0.001) but had no effect on wild-type cells (graph). Statistical analysis was performed using a two-sample Student’s t-Test with a two-tailed distribution. Standard deviation bars: n = 8.
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Fig6: Enhanced locomotion induced by tetracycline in T-REx-293 cells engineered withCrk-II.(a) DAPI staining (blue) and phalloidin-FITC (green) during scratch closing assays showing enhanced lamellipodia formation in THLOII-15 cells (a representative clone of T-REx-293 cells carrying the locomotion module) when induced with tetracycline: the induced cells show more frequent formation of lamellipodia, shown in green. Scale bars: 100 μm. (b) Distance travelled by cells in scratch closing assays with or without induction of the locomotion module. Images were acquired every 10 min and coordinates of 4 groups of cells on each side of wounds were recorded (6 to 18 h post-induction). Tetracycline increased the migration of THLOII-15 cells (trail plot and graph, *p <0.001) but had no effect on wild-type cells (graph). Statistical analysis was performed using a two-sample Student’s t-Test with a two-tailed distribution. Standard deviation bars: n = 8.

Mentions: A common way of studying cell locomotion is a scratch assay, in which a strip of cells is removed from a monolayer and the behaviour of remaining cells can be observed as they re-colonize the bare zone. Wild-type T-REx-293 cells (Additional file 2: Movie 2, Additional file 3: Movie 3) and T-REx-293 cells containing the locomotion module but with no induction (Additional file 4: Movie 4), showed only modest locomotory activity; some cells occasionally presenting a lamellipodium detectable by phalloidin staining for actin filaments (Figure 6a). When induced, cells expressing CRK-II exhibited enhanced formation of lamellipodia (Figure 6a, Additional file 5: Movie 5), repopulated the scratched area significantly faster and showed almost doubled motility (Figure 6b): the motility of CRK-II-expressing cells increased by a factor of 1.94, with p < 0.001 by Student’s t-Test. Wild-type T-REx-293 cells did not show enhanced motility in the presence of tetracycline.Figure 6


A library of mammalian effector modules for synthetic morphology.

Cachat E, Liu W, Hohenstein P, Davies JA - J Biol Eng (2014)

Enhanced locomotion induced by tetracycline in T-REx-293 cells engineered withCrk-II.(a) DAPI staining (blue) and phalloidin-FITC (green) during scratch closing assays showing enhanced lamellipodia formation in THLOII-15 cells (a representative clone of T-REx-293 cells carrying the locomotion module) when induced with tetracycline: the induced cells show more frequent formation of lamellipodia, shown in green. Scale bars: 100 μm. (b) Distance travelled by cells in scratch closing assays with or without induction of the locomotion module. Images were acquired every 10 min and coordinates of 4 groups of cells on each side of wounds were recorded (6 to 18 h post-induction). Tetracycline increased the migration of THLOII-15 cells (trail plot and graph, *p <0.001) but had no effect on wild-type cells (graph). Statistical analysis was performed using a two-sample Student’s t-Test with a two-tailed distribution. Standard deviation bars: n = 8.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4255936&req=5

Fig6: Enhanced locomotion induced by tetracycline in T-REx-293 cells engineered withCrk-II.(a) DAPI staining (blue) and phalloidin-FITC (green) during scratch closing assays showing enhanced lamellipodia formation in THLOII-15 cells (a representative clone of T-REx-293 cells carrying the locomotion module) when induced with tetracycline: the induced cells show more frequent formation of lamellipodia, shown in green. Scale bars: 100 μm. (b) Distance travelled by cells in scratch closing assays with or without induction of the locomotion module. Images were acquired every 10 min and coordinates of 4 groups of cells on each side of wounds were recorded (6 to 18 h post-induction). Tetracycline increased the migration of THLOII-15 cells (trail plot and graph, *p <0.001) but had no effect on wild-type cells (graph). Statistical analysis was performed using a two-sample Student’s t-Test with a two-tailed distribution. Standard deviation bars: n = 8.
Mentions: A common way of studying cell locomotion is a scratch assay, in which a strip of cells is removed from a monolayer and the behaviour of remaining cells can be observed as they re-colonize the bare zone. Wild-type T-REx-293 cells (Additional file 2: Movie 2, Additional file 3: Movie 3) and T-REx-293 cells containing the locomotion module but with no induction (Additional file 4: Movie 4), showed only modest locomotory activity; some cells occasionally presenting a lamellipodium detectable by phalloidin staining for actin filaments (Figure 6a). When induced, cells expressing CRK-II exhibited enhanced formation of lamellipodia (Figure 6a, Additional file 5: Movie 5), repopulated the scratched area significantly faster and showed almost doubled motility (Figure 6b): the motility of CRK-II-expressing cells increased by a factor of 1.94, with p < 0.001 by Student’s t-Test. Wild-type T-REx-293 cells did not show enhanced motility in the presence of tetracycline.Figure 6

Bottom Line: Together with cell differentiation, these mechanisms allow populations of cells to organize themselves into defined geometries and structures, as simple embryos develop into complex organisms.Here we describe this library and demonstrate its use in the T-REx-293 human cell line to induce each of these desired morphological behaviours on command.Building on from the simple test systems described here, we want to extend engineered control of morphogenetic cell behaviour to more complex 3D structures that can inform embryologists and may, in the future, be used in surgery and regenerative medicine, making synthetic morphology a powerful tool for developmental biology and tissue engineering.

View Article: PubMed Central - PubMed

Affiliation: University of Edinburgh, Centre for Integrative Physiology, Hugh Robson Building, George Square, Edinburgh, EH8 9XD UK.

ABSTRACT

Background: In mammalian development, the formation of most tissues is achieved by a relatively small repertoire of basic morphogenetic events (e.g. cell adhesion, locomotion, apoptosis, etc.), permutated in various sequences to form different tissues. Together with cell differentiation, these mechanisms allow populations of cells to organize themselves into defined geometries and structures, as simple embryos develop into complex organisms. The control of tissue morphogenesis by populations of engineered cells is a potentially very powerful but neglected aspect of synthetic biology.

Results: We have assembled a modular library of synthetic morphogenetic driver genes to control (separately) mammalian cell adhesion, locomotion, fusion, proliferation and elective cell death. Here we describe this library and demonstrate its use in the T-REx-293 human cell line to induce each of these desired morphological behaviours on command.

Conclusions: Building on from the simple test systems described here, we want to extend engineered control of morphogenetic cell behaviour to more complex 3D structures that can inform embryologists and may, in the future, be used in surgery and regenerative medicine, making synthetic morphology a powerful tool for developmental biology and tissue engineering.

No MeSH data available.


Related in: MedlinePlus