- Email: [email protected]
An Axonal Blueprint: Generating Neuronal Polarity with Light-Inducible Proteins
Cell Chemical Biology
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An Axonal Blueprint: Generating Neuronal Polarity with Light-Inducible Proteins Michael Z. Lin1,2,3,4,* 1Stanford
University Department of Neurobiology, Stanford, CA 94305, USA University Department of Bioengineering, Stanford, CA 94305, USA 3Stanford University Department of Chemical and Systems Biology, Stanford, CA 94305, USA 4Stanford University Department of Pediatrics, Stanford, CA 94305, USA *Correspondence: [email protected] https://doi.org/10.1016/j.chembiol.2019.12.003 2Stanford
In this issue of Cell Chemical Biology, Woo et al. (2019) show that activation of a photoinducible form of the TrkB protein in a single neurite induces multiple aspects of axonal differentiation. This study exemplifies the ability of optical methods to relate protein functions to complex phenotypes in living cells. Neurons are uniquely polarized cells, with an axon extending from the cell body to conduct action potentials, which in turn trigger neurotransmitter release and activation of downstream neurons. In the mammalian CNS, neurons extend multiple neurites, of which only one will develop into an axon. How a neuron selects one neurite to become an axon has been a long-standing topic of investigation in developmental neurobiology (Craig and Banker, 1994). In a study in this issue of Cell Chemical Biology, the group of Won Do Heo used a photoactivatable form of the TrkB receptor for brain-derived neurotrophic factor (BDNF), Opto-cytTrkB, to study how TrkB might promote axon growth and differentiation (Woo et al., 2019). OptocytTrkB consists of the myristylated cytosolic domain of TrkB fused to the PHR domain of Arabidopsis cryptochrome2 (CRY2PHR), which oligomerizes in response to blue light (Figure 1). Activation of Opto-cytTrkB by light applied to the tip of a neurite, but not light applied to neurite shafts, induced growth cone enlargement and actin wave propagation specifically in that neurite (Figure 1). These actin waves moved centrifugally from the cell body to the neurite tip over the course of about 30 min, and continued Opto-cytTrkB activation at neurite tips elicited additional waves (Figure 1). Woo et al. (2019) then showed by time-lapse microscopy that the stimulated neurite specifically accumulated fusions of a red fluorescent protein with ankyrinG, EB3, tau, drebrinE, or synaptophysin, proteins that mark axons in mature neurons (Figure 1). The local enrichment of these
protein reporters, despite their lacking control by native promoters or RNA untranslated regions, suggests accumulation was due to protein recruitment rather than local synthesis. Indeed, each of these markers could be seen decreasing in unstimulated neurites at the same time as they accumulated in stimulated neurites. The Rho-family GTPases Rac1 and Cdc42 can initiate actin polymerization via the Arp2/3 nucleating complex and are known to be activated downstream of the TrkB effector phosphatidylinositol 3-kinase (PI3K) (Lien et al., 2017). Thus, Rac1 and Cdc42 are prime candidates for mediating signals from TrkB to actin. Indeed, inhibition of Rac1 with a drug reduced the Opto-cytoTrkB-induced initiation of actin waves, but waves that did initiate still propagated centrifugally, suggesting Rac1 is required only for their triggering. Actin waves could be initiated by local illumination of a photoactivatable Rac1 protein at the soma but not at the axon tip, also consistent with a specific function for Rac1 in wave initiation. In contrast, a Cdc42 inhibitor caused OptocytoTrkB activation in a single neurite to initiate waves in multiple neurites, suggesting local Cdc42 activation may help promote wave propagation in the stimulated neurite and thereby direct the effects of Rac1 activation in the cell body away from other neurites. PI3K activity was also required for Opto-cytoTrkBinduced wave generation, consistent with its known ability to activate Cdc42 and Rac1. The time-lapse videos in the study of Woo et al. (2019) provide a vivid illustra-
1634 Cell Chemical Biology 26, December 19, 2019 ª 2019 Published by Elsevier Ltd.
tion of how waves alter the actin distribution in cells and hints at a solution for a chicken-and-egg problem in neurite extension. Neurites contain filamentous actin (F-actin) along their length, but this requires the continual presence of free globular actin (G-actin) to maintain, as F-actin is constantly in dynamic equilibrium with G-actin. Thus, a longer neurite should have proportionally more total actin than a shorter neurite of the same width. Interestingly, a mCherry-actin reporter revealed no increase in total cellular actin after optically induced neurite growth, suggesting Opto-cytTrkB was not stimulating actin synthesis. In addition, loss of total actin was apparent from non-stimulated neurites at the same time as the stimulated neurite was growing. How does actin get transported from non-growing neurites to the growing neurite? The answer could be the actin waves themselves, combined with diffusion. The assembly of G-actin at the plus end of an actin filament, which should point distally within the growthcone-like actin waves, could lead to a gradient of G-actin from high to low in the centrifugal direction. Diffusion of G-actin down this gradient could then serve to move G-actin centrifugally. Actin filaments can release G-actin at their negative ends to provide more subunits for recycling and incorporation into the distal plus ends. As long as retrograde movement of the F-actin does not exceed the rate of F-actin elongation by polymerization, the effect of a wave will be to continually shift the total actin pool centrifugally. As actin-GTP subunits have high affinity for plus ends, but are
Cell Chemical Biology
Previews
Figure 1. Local Induction of Actin Waves and Axonal Differentiation by Optical Activation of TrkB Signaling Opto-cytTrkB is a photo-inducible form of the cytosolic domain of TrkB (center). Activation of OptocytTrkB at a neurite tip by local blue light application (top left) induces an actin wave in the stimulated neurite. The wave is initiated at the cell body in a Rac1-dependent manner (top right) and propagated centrifugally along the stimulated neurite in a Cdc42-dependent manner (bottom right). Repeated activation of Opto-cytTrkB at the neurite tip induces additional actin waves, localization of axonal markers, and elongation of the stimulated neurite (bottom left).
converted to actin-GDP within the filament, which then shows enhanced dissociation at minus ends, energy for the process of directed diffusion may be supplied by GTP hydrolysis. Indeed, a function for actin waves in actin transport has been previously proposed (Inagaki and Katsuno, 2017). In this study, optical control enabled activation of Opto-cytTrkB only at the tips of specific neurites, an effect that would be possible to achieve by local perfusion of BDNF, but with much greater difficulty. Notably, earlier studies demonstrating BDNF-induced neuritic actin waves simply applied BDNF to the entire bath (Difato et al., 2011) and so could not study the role of BDNF in axonal specification, which requires stimulating one neurite only. One caveat of the current
experiments, and one that is common to all experiments involving the introduction of optically or chemically controllable proteins, is that the observed effects are those of exogenous proteins and thus may not fully reflect the functions of endogenous proteins. Thus, the relationship of observed phenotypes to native protein functions can only be established by studying unmodified cells or cells with loss-of-function perturbations of native proteins (Polleux and Snider, 2010). The function of BDNF in axon extension has already been established in this way; e.g., the first neurite to contact immobilized BDNF develops into an axon, and anti-BDNF antibodies reduce the formation of axons (Cheng and Poo, 2012; Polleux and Snider, 2010). Thus, the observed mechanisms by which Opto-cytTrkB
induce axon growth and differentiation are likely to be physiologically relevant. Remaining questions relate to how exactly TrkB induces axonal differentiation. For example, does TrkB activate signals previously implicated in axonal maturation, or does it promote effective axonal differentiation independently of some of these signals, or does it only induce a subset of the axonal phenotype? For example, whether TrkB is capable of relocalizing the PAR3/6 complex and activating LKB1, events previously found to be required for axon formation (Polleux and Snider, 2010), would be useful to know. The speed at which ankyrinG relocalizes to the activated neurite (within the 2 h of experimental observation) suggests rapid and direct involvement of TrkB effectors in regulating ankyrinG localization without requiring slower processes such as transcriptional programs of cell differentiation. That is, the axon initial segment, which contains ankyrinG, is not completely a late structure; rather, its main structural component, ankyrinG, can be recruited to one neurite simultaneously with initial establishment of axon-dendrite polarity. This positions ankyrinG to potentially support other morphological events in axonal differentiation, such as the polarity sorting of microtubules. Finally, can we test the hypothesis that actin waves are required for actin transport to the axon to enable further extension? If transport of actin is an essential outcome, then it might be expected that stopping waves would stop further neurite lengthening. While Rac inhibitors can suppress wave initiation, they can also be expected to block growth cone extension at the neurite tip, so a phenotype of reduced axonal extension might not be due to wave suppression alone. With the recent development of Z-lock cofilin, a photoactivatable form of the actin-severing protein cofilin (Stone et al., 2019), stopping waves only in the middle of neurites is conceivably possible. However, as both OptocytTrkB and Z-lock cofilin use flavin chromophores that absorb blue light, they cannot be used orthogonally within the same experiment. Designing a photoactivatable cofilin to use a different wavelength, such as by replacing the Z-lock domains with cyan-absorbing pdDronpa
Cell Chemical Biology 26, December 19, 2019 1635
Cell Chemical Biology
Previews
domains (Zhou et al., 2017), may be helpful in this regard. In summary, the article by Woo et al. (2019) demonstrates the power of photoregulation of proteins to investigate protein function in living cells with exquisite spatiotemporal precision. Since the first use of photosensory domains to control protein activities in 2002, the number of studies using optobiological approaches to investigate dynamic aspects of protein signaling in living cells has steadily increased (Goglia and Toettcher, 2019). The article by Woo et al. serves as an exciting example of how classic cell biology questions can be re-investigated using modern methods, resulting in new discoveries of underlying molecular mechanisms.
REFERENCES Cheng, P.L., and Poo, M.M. (2012). Early events in axon/dendrite polarization. Annu. Rev. Neurosci. 35, 181–201. Craig, A.M., and Banker, G. (1994). Neuronal polarity. Annu. Rev. Neurosci. 17, 267–310. Difato, F., Tsushima, H., Pesce, M., Benfenati, F., Blau, A., and Chieregatti, E. (2011). The formation of actin waves during regeneration after axonal lesion is enhanced by BDNF. Sci. Rep. 1, 183. Goglia, A.G., and Toettcher, J.E. (2019). A bright future: optogenetics to dissect the spatiotemporal control of cell behavior. Curr. Opin. Chem. Biol. 48, 106–113. Inagaki, N., and Katsuno, H. (2017). Actin Waves: Origin of Cell Polarization and Migration? Trends Cell Biol. 27, 515–526.
Lien, E.C., Dibble, C.C., and Toker, A. (2017). PI3K signaling in cancer: beyond AKT. Curr. Opin. Cell Biol. 45, 62–71. Polleux, F., and Snider, W. (2010). Initiating and growing an axon. Cold Spring Harb. Perspect. Biol. 2, a001925. Stone, O.J., Pankow, N., Liu, B., Sharma, V.P., Eddy, R.J., Wang, H., Putz, A.T., Teets, F.D., Kuhlman, B., Condeelis, J.S., and Hahn, K.M. (2019). Optogenetic control of cofilin and aTAT in living cells using Z-lock. Nat. Chem. Biol. 15, 1183–1190. Woo, D., Seo, Y., Jung, H., Kim, S., Kim, N., Park, S.M., Lee, H., Lee, S., Cho, K.H., and Heo, W.D. (2019). Locally activating TrkB receptor generates actin waves and specifies axonal fate. Cell Chem. Biol. 26, this issue, 1652–1663. Zhou, X.X., Fan, L.Z., Li, P., Shen, K., and Lin, M.Z. (2017). Optical control of cell signaling by singlechain photoswitchable kinases. Science 355, 836–842.
Death by Retrograde Transport: Avoiding the Apoptosis Default Douglas R. Green1,* 1Department of Immunology, St. Jude Children’s Research Institute, Memphis, TN 38105, USA *Correspondence: [email protected] https://doi.org/10.1016/j.chembiol.2019.12.002
In this issue of Cell Chemical Biology, Ko et al. (2019) describe a form of regulated necrosis that depends on active, retrograde transport of vesicles from the plasma membrane to the Golgi. The cell death is distinct from defined modalities including apoptosis, and most tumor lines show enhanced sensitivity to it. Just as there are many ways to live, there are a great many ways that cells can die. Cell death can be passive, akin to ‘‘accident’’ or ‘‘murder,’’ where cells are simply damaged beyond repair. Perhaps more intriguing (and informative) are situations where cells actively participate in their own demise. Such cell death can be a form of ‘‘suicide,’’ involving pathways dedicated to killing the cell, including apoptosis, necroptosis, and pyroptosis (Green, 2018). Alternatively, insults such as chemical agents can effectively ‘‘sabotage’’ active cellular processes that sustain cell survival, such that cells die only when such processes are active and required for life. Examples of the latter include ferroptosis, where mechanisms to prevent lethal lipid peroxidation are de-
railed, and parthanatos, where the excessive engagement of DNA repair pathways (i.e., Poly-ADP ribose polymerase) depletes essential energy stores (Green, 2018). In this issue, Dixon and colleagues (Ko et al., 2019) describe a chemical agent, CIL56, that preferentially kills many tumor cell lines while sparing normal cells, by sabotaging a transport process, leading to death of the cell via a modality that appears to be distinct from any so far described. Inhibitors of apoptosis, necroptosis, pyroptosis, or ferroptosis had no effect on cell death induced by CIL56. CIL56 was identified in a screen for compounds capable of preferentially killing tumor lines (Dixon et al., 2015). The authors now find that of 94 cancer lines examined, CIL56 potently kills
1636 Cell Chemical Biology 26, December 19, 2019 ª 2019 Published by Elsevier Ltd.
most hematologic cancer lines, as well as many others (although, notably, not those of liver or muscle origin), while completely sparing normal peripheral blood mononuclear cells (PBMCs) (Ko et al., 2019). Consistent with their earlier findings (Dixon et al., 2015), and using an siRNA screen, they have now confirmed that toxicity of CIL56 depends on fatty acid synthesis, as silencing or pharmacologic inhibition of fatty acid synthase (Fasn), acetyl-CoA carboxylase-1 (ACC1), or long-chain-fattyacid-CoA ligase-3 (ACSL3) all prevent killing by this agent. Of particular note, however, was the identification of two other requisite proteins, Zinc finger DHHC domain-containing protein-5 (ZDHHC5) and Golgin subfamily A member-7 (Golga7), both of which were
Previews
An Axonal Blueprint: Generating Neuronal Polarity with Light-Inducible Proteins Michael Z. Lin1,2,3,4,* 1Stanford
University Department of Neurobiology, Stanford, CA 94305, USA University Department of Bioengineering, Stanford, CA 94305, USA 3Stanford University Department of Chemical and Systems Biology, Stanford, CA 94305, USA 4Stanford University Department of Pediatrics, Stanford, CA 94305, USA *Correspondence: [email protected] https://doi.org/10.1016/j.chembiol.2019.12.003 2Stanford
In this issue of Cell Chemical Biology, Woo et al. (2019) show that activation of a photoinducible form of the TrkB protein in a single neurite induces multiple aspects of axonal differentiation. This study exemplifies the ability of optical methods to relate protein functions to complex phenotypes in living cells. Neurons are uniquely polarized cells, with an axon extending from the cell body to conduct action potentials, which in turn trigger neurotransmitter release and activation of downstream neurons. In the mammalian CNS, neurons extend multiple neurites, of which only one will develop into an axon. How a neuron selects one neurite to become an axon has been a long-standing topic of investigation in developmental neurobiology (Craig and Banker, 1994). In a study in this issue of Cell Chemical Biology, the group of Won Do Heo used a photoactivatable form of the TrkB receptor for brain-derived neurotrophic factor (BDNF), Opto-cytTrkB, to study how TrkB might promote axon growth and differentiation (Woo et al., 2019). OptocytTrkB consists of the myristylated cytosolic domain of TrkB fused to the PHR domain of Arabidopsis cryptochrome2 (CRY2PHR), which oligomerizes in response to blue light (Figure 1). Activation of Opto-cytTrkB by light applied to the tip of a neurite, but not light applied to neurite shafts, induced growth cone enlargement and actin wave propagation specifically in that neurite (Figure 1). These actin waves moved centrifugally from the cell body to the neurite tip over the course of about 30 min, and continued Opto-cytTrkB activation at neurite tips elicited additional waves (Figure 1). Woo et al. (2019) then showed by time-lapse microscopy that the stimulated neurite specifically accumulated fusions of a red fluorescent protein with ankyrinG, EB3, tau, drebrinE, or synaptophysin, proteins that mark axons in mature neurons (Figure 1). The local enrichment of these
protein reporters, despite their lacking control by native promoters or RNA untranslated regions, suggests accumulation was due to protein recruitment rather than local synthesis. Indeed, each of these markers could be seen decreasing in unstimulated neurites at the same time as they accumulated in stimulated neurites. The Rho-family GTPases Rac1 and Cdc42 can initiate actin polymerization via the Arp2/3 nucleating complex and are known to be activated downstream of the TrkB effector phosphatidylinositol 3-kinase (PI3K) (Lien et al., 2017). Thus, Rac1 and Cdc42 are prime candidates for mediating signals from TrkB to actin. Indeed, inhibition of Rac1 with a drug reduced the Opto-cytoTrkB-induced initiation of actin waves, but waves that did initiate still propagated centrifugally, suggesting Rac1 is required only for their triggering. Actin waves could be initiated by local illumination of a photoactivatable Rac1 protein at the soma but not at the axon tip, also consistent with a specific function for Rac1 in wave initiation. In contrast, a Cdc42 inhibitor caused OptocytoTrkB activation in a single neurite to initiate waves in multiple neurites, suggesting local Cdc42 activation may help promote wave propagation in the stimulated neurite and thereby direct the effects of Rac1 activation in the cell body away from other neurites. PI3K activity was also required for Opto-cytoTrkBinduced wave generation, consistent with its known ability to activate Cdc42 and Rac1. The time-lapse videos in the study of Woo et al. (2019) provide a vivid illustra-
1634 Cell Chemical Biology 26, December 19, 2019 ª 2019 Published by Elsevier Ltd.
tion of how waves alter the actin distribution in cells and hints at a solution for a chicken-and-egg problem in neurite extension. Neurites contain filamentous actin (F-actin) along their length, but this requires the continual presence of free globular actin (G-actin) to maintain, as F-actin is constantly in dynamic equilibrium with G-actin. Thus, a longer neurite should have proportionally more total actin than a shorter neurite of the same width. Interestingly, a mCherry-actin reporter revealed no increase in total cellular actin after optically induced neurite growth, suggesting Opto-cytTrkB was not stimulating actin synthesis. In addition, loss of total actin was apparent from non-stimulated neurites at the same time as the stimulated neurite was growing. How does actin get transported from non-growing neurites to the growing neurite? The answer could be the actin waves themselves, combined with diffusion. The assembly of G-actin at the plus end of an actin filament, which should point distally within the growthcone-like actin waves, could lead to a gradient of G-actin from high to low in the centrifugal direction. Diffusion of G-actin down this gradient could then serve to move G-actin centrifugally. Actin filaments can release G-actin at their negative ends to provide more subunits for recycling and incorporation into the distal plus ends. As long as retrograde movement of the F-actin does not exceed the rate of F-actin elongation by polymerization, the effect of a wave will be to continually shift the total actin pool centrifugally. As actin-GTP subunits have high affinity for plus ends, but are
Cell Chemical Biology
Previews
Figure 1. Local Induction of Actin Waves and Axonal Differentiation by Optical Activation of TrkB Signaling Opto-cytTrkB is a photo-inducible form of the cytosolic domain of TrkB (center). Activation of OptocytTrkB at a neurite tip by local blue light application (top left) induces an actin wave in the stimulated neurite. The wave is initiated at the cell body in a Rac1-dependent manner (top right) and propagated centrifugally along the stimulated neurite in a Cdc42-dependent manner (bottom right). Repeated activation of Opto-cytTrkB at the neurite tip induces additional actin waves, localization of axonal markers, and elongation of the stimulated neurite (bottom left).
converted to actin-GDP within the filament, which then shows enhanced dissociation at minus ends, energy for the process of directed diffusion may be supplied by GTP hydrolysis. Indeed, a function for actin waves in actin transport has been previously proposed (Inagaki and Katsuno, 2017). In this study, optical control enabled activation of Opto-cytTrkB only at the tips of specific neurites, an effect that would be possible to achieve by local perfusion of BDNF, but with much greater difficulty. Notably, earlier studies demonstrating BDNF-induced neuritic actin waves simply applied BDNF to the entire bath (Difato et al., 2011) and so could not study the role of BDNF in axonal specification, which requires stimulating one neurite only. One caveat of the current
experiments, and one that is common to all experiments involving the introduction of optically or chemically controllable proteins, is that the observed effects are those of exogenous proteins and thus may not fully reflect the functions of endogenous proteins. Thus, the relationship of observed phenotypes to native protein functions can only be established by studying unmodified cells or cells with loss-of-function perturbations of native proteins (Polleux and Snider, 2010). The function of BDNF in axon extension has already been established in this way; e.g., the first neurite to contact immobilized BDNF develops into an axon, and anti-BDNF antibodies reduce the formation of axons (Cheng and Poo, 2012; Polleux and Snider, 2010). Thus, the observed mechanisms by which Opto-cytTrkB
induce axon growth and differentiation are likely to be physiologically relevant. Remaining questions relate to how exactly TrkB induces axonal differentiation. For example, does TrkB activate signals previously implicated in axonal maturation, or does it promote effective axonal differentiation independently of some of these signals, or does it only induce a subset of the axonal phenotype? For example, whether TrkB is capable of relocalizing the PAR3/6 complex and activating LKB1, events previously found to be required for axon formation (Polleux and Snider, 2010), would be useful to know. The speed at which ankyrinG relocalizes to the activated neurite (within the 2 h of experimental observation) suggests rapid and direct involvement of TrkB effectors in regulating ankyrinG localization without requiring slower processes such as transcriptional programs of cell differentiation. That is, the axon initial segment, which contains ankyrinG, is not completely a late structure; rather, its main structural component, ankyrinG, can be recruited to one neurite simultaneously with initial establishment of axon-dendrite polarity. This positions ankyrinG to potentially support other morphological events in axonal differentiation, such as the polarity sorting of microtubules. Finally, can we test the hypothesis that actin waves are required for actin transport to the axon to enable further extension? If transport of actin is an essential outcome, then it might be expected that stopping waves would stop further neurite lengthening. While Rac inhibitors can suppress wave initiation, they can also be expected to block growth cone extension at the neurite tip, so a phenotype of reduced axonal extension might not be due to wave suppression alone. With the recent development of Z-lock cofilin, a photoactivatable form of the actin-severing protein cofilin (Stone et al., 2019), stopping waves only in the middle of neurites is conceivably possible. However, as both OptocytTrkB and Z-lock cofilin use flavin chromophores that absorb blue light, they cannot be used orthogonally within the same experiment. Designing a photoactivatable cofilin to use a different wavelength, such as by replacing the Z-lock domains with cyan-absorbing pdDronpa
Cell Chemical Biology 26, December 19, 2019 1635
Cell Chemical Biology
Previews
domains (Zhou et al., 2017), may be helpful in this regard. In summary, the article by Woo et al. (2019) demonstrates the power of photoregulation of proteins to investigate protein function in living cells with exquisite spatiotemporal precision. Since the first use of photosensory domains to control protein activities in 2002, the number of studies using optobiological approaches to investigate dynamic aspects of protein signaling in living cells has steadily increased (Goglia and Toettcher, 2019). The article by Woo et al. serves as an exciting example of how classic cell biology questions can be re-investigated using modern methods, resulting in new discoveries of underlying molecular mechanisms.
REFERENCES Cheng, P.L., and Poo, M.M. (2012). Early events in axon/dendrite polarization. Annu. Rev. Neurosci. 35, 181–201. Craig, A.M., and Banker, G. (1994). Neuronal polarity. Annu. Rev. Neurosci. 17, 267–310. Difato, F., Tsushima, H., Pesce, M., Benfenati, F., Blau, A., and Chieregatti, E. (2011). The formation of actin waves during regeneration after axonal lesion is enhanced by BDNF. Sci. Rep. 1, 183. Goglia, A.G., and Toettcher, J.E. (2019). A bright future: optogenetics to dissect the spatiotemporal control of cell behavior. Curr. Opin. Chem. Biol. 48, 106–113. Inagaki, N., and Katsuno, H. (2017). Actin Waves: Origin of Cell Polarization and Migration? Trends Cell Biol. 27, 515–526.
Lien, E.C., Dibble, C.C., and Toker, A. (2017). PI3K signaling in cancer: beyond AKT. Curr. Opin. Cell Biol. 45, 62–71. Polleux, F., and Snider, W. (2010). Initiating and growing an axon. Cold Spring Harb. Perspect. Biol. 2, a001925. Stone, O.J., Pankow, N., Liu, B., Sharma, V.P., Eddy, R.J., Wang, H., Putz, A.T., Teets, F.D., Kuhlman, B., Condeelis, J.S., and Hahn, K.M. (2019). Optogenetic control of cofilin and aTAT in living cells using Z-lock. Nat. Chem. Biol. 15, 1183–1190. Woo, D., Seo, Y., Jung, H., Kim, S., Kim, N., Park, S.M., Lee, H., Lee, S., Cho, K.H., and Heo, W.D. (2019). Locally activating TrkB receptor generates actin waves and specifies axonal fate. Cell Chem. Biol. 26, this issue, 1652–1663. Zhou, X.X., Fan, L.Z., Li, P., Shen, K., and Lin, M.Z. (2017). Optical control of cell signaling by singlechain photoswitchable kinases. Science 355, 836–842.
Death by Retrograde Transport: Avoiding the Apoptosis Default Douglas R. Green1,* 1Department of Immunology, St. Jude Children’s Research Institute, Memphis, TN 38105, USA *Correspondence: [email protected] https://doi.org/10.1016/j.chembiol.2019.12.002
In this issue of Cell Chemical Biology, Ko et al. (2019) describe a form of regulated necrosis that depends on active, retrograde transport of vesicles from the plasma membrane to the Golgi. The cell death is distinct from defined modalities including apoptosis, and most tumor lines show enhanced sensitivity to it. Just as there are many ways to live, there are a great many ways that cells can die. Cell death can be passive, akin to ‘‘accident’’ or ‘‘murder,’’ where cells are simply damaged beyond repair. Perhaps more intriguing (and informative) are situations where cells actively participate in their own demise. Such cell death can be a form of ‘‘suicide,’’ involving pathways dedicated to killing the cell, including apoptosis, necroptosis, and pyroptosis (Green, 2018). Alternatively, insults such as chemical agents can effectively ‘‘sabotage’’ active cellular processes that sustain cell survival, such that cells die only when such processes are active and required for life. Examples of the latter include ferroptosis, where mechanisms to prevent lethal lipid peroxidation are de-
railed, and parthanatos, where the excessive engagement of DNA repair pathways (i.e., Poly-ADP ribose polymerase) depletes essential energy stores (Green, 2018). In this issue, Dixon and colleagues (Ko et al., 2019) describe a chemical agent, CIL56, that preferentially kills many tumor cell lines while sparing normal cells, by sabotaging a transport process, leading to death of the cell via a modality that appears to be distinct from any so far described. Inhibitors of apoptosis, necroptosis, pyroptosis, or ferroptosis had no effect on cell death induced by CIL56. CIL56 was identified in a screen for compounds capable of preferentially killing tumor lines (Dixon et al., 2015). The authors now find that of 94 cancer lines examined, CIL56 potently kills
1636 Cell Chemical Biology 26, December 19, 2019 ª 2019 Published by Elsevier Ltd.
most hematologic cancer lines, as well as many others (although, notably, not those of liver or muscle origin), while completely sparing normal peripheral blood mononuclear cells (PBMCs) (Ko et al., 2019). Consistent with their earlier findings (Dixon et al., 2015), and using an siRNA screen, they have now confirmed that toxicity of CIL56 depends on fatty acid synthesis, as silencing or pharmacologic inhibition of fatty acid synthase (Fasn), acetyl-CoA carboxylase-1 (ACC1), or long-chain-fattyacid-CoA ligase-3 (ACSL3) all prevent killing by this agent. Of particular note, however, was the identification of two other requisite proteins, Zinc finger DHHC domain-containing protein-5 (ZDHHC5) and Golgin subfamily A member-7 (Golga7), both of which were