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Making axons with dendrites
448
IIt~[~v~v~,'~,~'l[l~Jziill|[~tkll
TRENDS in Cell Biology Vol.11 N o . l l N o v e m b e r 2001
Pushing the envelope in mitosis One of the first events of mitosis is nuclear envelope (NE) breakdown. During prophase, the NE dissociates into vesicles that disperse t h r o u g h o u t the cell. Recent work, however, suggests that a NE-associated protein plays an i m p o r t a n t role in c h r o m o s o m e segregation in Drosophila, in spite of the fact that the N E is not present during mitosis. The Su(var)2-1Ogene of Drosophila was first identified because mutants in this gene are defective in position effect variegation (PEV), a process in which genes are silenced by nearby heterochromatin. Karpen and c o w o r k e r s ~ found that Su(var)2-10 mutations also showed defects in m i n i c h r o m o s o m e transmission, suggesting that this gene plays a role in c h r o m o s o m e segregation. Cytological analysis of m u t a n t animals lacking w i l d - t y p e Su(var)2-10 revealed that mitosis is highly aberrant. In metaphase, c h r o m o s o m e s are large and puffy, indicating a defect in c h r o m o s o m e condensation. In anaphase, the Su(var)2-10 mutation caused c h r o m o s o m e bridges and fragmentation. Given that Su(var)2-10 mutations cause defects in a heterochromatin-based process (PEV) as well as c h r o m o s o m e condensation and
segregation, the obvious guess w o u l d be that it encodes a c h r o m o s o m a l protein. Indeed, cloning and sequencing revealed that Su(var)2-1Ocontained potential DNAbinding motifs. Surprisingly, i m m u n o f l u o r e s c e n c e showed that Su(var)2-10 does not localize to mitotic chromosomes. Instead, during interphase, the protein is concentrated mainly at the nuclear envelope, colocalizing w i t h nuclear lamins. Although there are also some intranuclear loci, these do not generally colocalize with chromatin. If w e take these data at face value and conclude that the p r i m a r y site of Su(var)2-10 function is the nuclear envelope, w e are left w i t h an interesting puzzle: h o w does a protein localized on the nuclear envelope influence so many different aspects of c h r o m o s o m e behavior? A possible solution to this d i l e m m a came from e x a m i n i n g the interphase nuclear architecture in Su(var)2-10 mutants. Fluorescence in situ hybridization (FISH) using t e l o m e r i c probes, revealed that the t e l o m e r e clustering seen in normal cells, as well as the normal association of t e l o m e r e s with the nuclear envelope, was disrupted in the
mutants. Thus, Su(var)2-10 m i g h t play a role in mediating interactions between specific c h r o m o s o m a l regions, such as telomeres, with the nuclear envelope. Defects in Su(var)2-1Ocould therefore have far-reaching consequences for p h e n o m e n a such as PEV that involve long-range interactions w i t h i n the nucleus and are thus sensitive to changes in nuclear organization. But w h a t about mitosis? Mitotic c h r o m o s o m e condensation in Drosophila initiates at a small n u m b e r of condensation foci that are always associated with the nuclear envelope. If Su(var)2-10 plays a role in this interaction, m u t a t i o n s in the protein m i g h t interfere with the early stages of prophase c h r o m o s o m e condensation, leading to defects that only become apparent later in mitosis. 1 Hari, K.L. etal, (2001) The Drosophila Su(var)2-10 locus regulates chromosome structure and function and encodes a member of the PIAS protein family. Genes Dev. 15,
1334-1348
Wallace E Marshall wfm5@ pantheon.yale.edu
Making axons with dendrites One of the fundamental questions of n e u r o b i o l o g y concerns the mysterious capacity of neurons to extend one axon and several dendrites. Such extreme control of cell polarity is progressively acquired during development. Current models suggest that, at least in culture, after a s y m m e t r i c o u t g r o w t h of several processes extending f r o m the cell body, positive- and negative-feedback regulation breakthe s y m m e t r y and favor the extension of a long and thin process the axon. This p h e n o m e n o n is intimately linked to modifications of the actin networks, a l l o w i n g process extension controlled by transient intracellular Ca 2+ elevations and activation of the GTPase Rho. Nevertheless, little is known about the molecular nature of the actors controlling the acquisition of neuronal polarity. KaibuchP and colleagues have n o w identified the essential r e q u i r e m e n t
for a cytosolic p h o s p h o p r o t e i n (CRMP-2 coliapsin response m e d i a t o r protein, also t e r m e d TOAD-64/Ulip-2/DRP-2) previously known to be involved in neuronal differentiation and axonal guidance. In cultures of h i p p o c a m p a l neurons, the authors found that, once a clear neuronal polarity is observed, high levels of CRMP-2 are detected in the g r o w t h cone and the distal part of the axon, whereas cell body and dendrites present l o w immunoreactivity. Strikingly, overexpression of CRMP-2 in polarized neurons triggers the f o r m a t i o n of additional axons. Using fluorescent EGFPtagged CRMP-2, they d e m o n s t r a t e d that increased levels of CRMP-2 confer an axonal phenotype not only to small e m e r g i n g neurites but also to established dendrites. Additional e x p e r i m e n t s in which truncated f o r m s of CRMP-2 were
transfected provided further evidence for a specific i n v o l v e m e n t of a C-terminal 24-residue f r a g m e n t during this acquisition of s u p e r n u m e r a r y axons. Overall, this study provides a powerful model to analyze the upstream and d o w n s t r e a m targets participating in the acquisition of neuronal polarity. The previous links already described between CRMP and axonal guidance (such as semaphorins), cytoskeleton dynamics and Rho-kinase herald a cascade of evidence that should n o w lead to a better comprehension of h o w to make a single axon. 1 Inagaki, N. etal. (2001) CRMP-2 induces axons in cultured hippocampal neurons. Nat. Neurosci. 4, 781-782
Dominique Bagnard bagnard @neu rochem.u-strasbg.fr
http://tcb.trends.com 0962-8924/01/$- see front matter © 2001 Elsevier Science Ltd. All rights reserved,
IIt~[~v~v~,'~,~'l[l~Jziill|[~tkll
TRENDS in Cell Biology Vol.11 N o . l l N o v e m b e r 2001
Pushing the envelope in mitosis One of the first events of mitosis is nuclear envelope (NE) breakdown. During prophase, the NE dissociates into vesicles that disperse t h r o u g h o u t the cell. Recent work, however, suggests that a NE-associated protein plays an i m p o r t a n t role in c h r o m o s o m e segregation in Drosophila, in spite of the fact that the N E is not present during mitosis. The Su(var)2-1Ogene of Drosophila was first identified because mutants in this gene are defective in position effect variegation (PEV), a process in which genes are silenced by nearby heterochromatin. Karpen and c o w o r k e r s ~ found that Su(var)2-10 mutations also showed defects in m i n i c h r o m o s o m e transmission, suggesting that this gene plays a role in c h r o m o s o m e segregation. Cytological analysis of m u t a n t animals lacking w i l d - t y p e Su(var)2-10 revealed that mitosis is highly aberrant. In metaphase, c h r o m o s o m e s are large and puffy, indicating a defect in c h r o m o s o m e condensation. In anaphase, the Su(var)2-10 mutation caused c h r o m o s o m e bridges and fragmentation. Given that Su(var)2-10 mutations cause defects in a heterochromatin-based process (PEV) as well as c h r o m o s o m e condensation and
segregation, the obvious guess w o u l d be that it encodes a c h r o m o s o m a l protein. Indeed, cloning and sequencing revealed that Su(var)2-1Ocontained potential DNAbinding motifs. Surprisingly, i m m u n o f l u o r e s c e n c e showed that Su(var)2-10 does not localize to mitotic chromosomes. Instead, during interphase, the protein is concentrated mainly at the nuclear envelope, colocalizing w i t h nuclear lamins. Although there are also some intranuclear loci, these do not generally colocalize with chromatin. If w e take these data at face value and conclude that the p r i m a r y site of Su(var)2-10 function is the nuclear envelope, w e are left w i t h an interesting puzzle: h o w does a protein localized on the nuclear envelope influence so many different aspects of c h r o m o s o m e behavior? A possible solution to this d i l e m m a came from e x a m i n i n g the interphase nuclear architecture in Su(var)2-10 mutants. Fluorescence in situ hybridization (FISH) using t e l o m e r i c probes, revealed that the t e l o m e r e clustering seen in normal cells, as well as the normal association of t e l o m e r e s with the nuclear envelope, was disrupted in the
mutants. Thus, Su(var)2-10 m i g h t play a role in mediating interactions between specific c h r o m o s o m a l regions, such as telomeres, with the nuclear envelope. Defects in Su(var)2-1Ocould therefore have far-reaching consequences for p h e n o m e n a such as PEV that involve long-range interactions w i t h i n the nucleus and are thus sensitive to changes in nuclear organization. But w h a t about mitosis? Mitotic c h r o m o s o m e condensation in Drosophila initiates at a small n u m b e r of condensation foci that are always associated with the nuclear envelope. If Su(var)2-10 plays a role in this interaction, m u t a t i o n s in the protein m i g h t interfere with the early stages of prophase c h r o m o s o m e condensation, leading to defects that only become apparent later in mitosis. 1 Hari, K.L. etal, (2001) The Drosophila Su(var)2-10 locus regulates chromosome structure and function and encodes a member of the PIAS protein family. Genes Dev. 15,
1334-1348
Wallace E Marshall wfm5@ pantheon.yale.edu
Making axons with dendrites One of the fundamental questions of n e u r o b i o l o g y concerns the mysterious capacity of neurons to extend one axon and several dendrites. Such extreme control of cell polarity is progressively acquired during development. Current models suggest that, at least in culture, after a s y m m e t r i c o u t g r o w t h of several processes extending f r o m the cell body, positive- and negative-feedback regulation breakthe s y m m e t r y and favor the extension of a long and thin process the axon. This p h e n o m e n o n is intimately linked to modifications of the actin networks, a l l o w i n g process extension controlled by transient intracellular Ca 2+ elevations and activation of the GTPase Rho. Nevertheless, little is known about the molecular nature of the actors controlling the acquisition of neuronal polarity. KaibuchP and colleagues have n o w identified the essential r e q u i r e m e n t
for a cytosolic p h o s p h o p r o t e i n (CRMP-2 coliapsin response m e d i a t o r protein, also t e r m e d TOAD-64/Ulip-2/DRP-2) previously known to be involved in neuronal differentiation and axonal guidance. In cultures of h i p p o c a m p a l neurons, the authors found that, once a clear neuronal polarity is observed, high levels of CRMP-2 are detected in the g r o w t h cone and the distal part of the axon, whereas cell body and dendrites present l o w immunoreactivity. Strikingly, overexpression of CRMP-2 in polarized neurons triggers the f o r m a t i o n of additional axons. Using fluorescent EGFPtagged CRMP-2, they d e m o n s t r a t e d that increased levels of CRMP-2 confer an axonal phenotype not only to small e m e r g i n g neurites but also to established dendrites. Additional e x p e r i m e n t s in which truncated f o r m s of CRMP-2 were
transfected provided further evidence for a specific i n v o l v e m e n t of a C-terminal 24-residue f r a g m e n t during this acquisition of s u p e r n u m e r a r y axons. Overall, this study provides a powerful model to analyze the upstream and d o w n s t r e a m targets participating in the acquisition of neuronal polarity. The previous links already described between CRMP and axonal guidance (such as semaphorins), cytoskeleton dynamics and Rho-kinase herald a cascade of evidence that should n o w lead to a better comprehension of h o w to make a single axon. 1 Inagaki, N. etal. (2001) CRMP-2 induces axons in cultured hippocampal neurons. Nat. Neurosci. 4, 781-782
Dominique Bagnard bagnard @neu rochem.u-strasbg.fr
http://tcb.trends.com 0962-8924/01/$- see front matter © 2001 Elsevier Science Ltd. All rights reserved,