Department of Biology, Massachusetts Institute of Technology, 77 Massachusetts Avenue 68-230B, Cambridge, MA 02139, USA
* Author for correspondence (e-mail: pgarrity{at}mit.edu)
Accepted 8 May 2003
![]() |
SUMMARY |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Key words: Pvr, Engulfment, Hemocyte, Cell death, Apoptosis, Drosophila
![]() |
INTRODUCTION |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Macrophages are responsible for the majority of cell corpse removal during
mammalian development (Hopkinson-Woolley
et al., 1994; Hume et al.,
1983
; Morris et al.,
1991
). In Drosophila, cell corpse removal also relies on
specialized phagocytic cells that resemble mammalian macrophages in a number
of cellular and molecular properties
(Franc, 2002
).
Drosophila macrophages are derived from hematopoietic precursor cells
known as hemocytes, which differentiate into macrophages displaying phagocytic
and scavenger properties in response to cell corpse exposure
(Tepass et al., 1994
). Work on
mammalian macrophages and other phagocytic cells has identified several
classes of receptors implicated in corpse recognition, including lectins,
integrins, the MER tyrosine kinase, the phosphatidylserine receptor (PSR) and
scavenger receptors such as CD36 (Fadok
and Chimini, 2001
). Drosophila macrophages express a
CD36-related receptor, Croquemort (Franc
et al., 1996
), and croquemort (crq) function is
required for macrophages to take up dead cells efficiently in the developing
embryo (Franc et al.,
1999
).
As macrophages are responsible for much of the dead cell engulfment in
developing animals, an important role for macrophages in tissue morphogenesis
during development has been suggested
(Morris et al., 1991).
However, direct evidence of a required role for macrophage-mediated cell
corpse engulfment in development is limited. In the feet of PU.1 mutant mice
that lack macrophages, for example, other cell types take over the engulfing
role and permit morphogenesis to proceed, albeit at a slower pace
(Wood et al., 2000
). By
contrast, in the developing mouse retina macrophages are essential for cell
death-mediated morphogenesis (Lang et al.,
1994
; Lang and Bishop,
1993
). In this case, the primary defect is not caused by a failure
of engulfment. Rather, macrophages are required to initiate the cell deaths
that normally eliminate the hyaloid vessels and the pupillary membrane during
the development of the mouse eye.
In the CNS of Drosophila melanogaster embryos, programmed cell
death eliminates many neurons and glia
(Jacobs, 2000;
Sonnenfeld and Jacobs, 1995b
).
In the case of the midline glia, approximately ten midline glial cells are
generated in each segment by stage 13 of embryonic development. As development
proceeds, most of these glia are eliminated by programmed cell death, leaving
two to three midline glia per segment by stage 17
(Klambt et al., 1991
;
Sonnenfeld and Jacobs, 1995a
;
Zhou et al., 1995
). Recent
work indicates that midline glial cell survival is mediated through activation
of MAP kinase signaling in the midline glia via the reception of the EGFR
ligand Spitz, which is provided by the developing neurons
(Bergmann et al., 2002
). In
addition to midline glia, a subset of developing neurons and longitudinal glia
(which flank the midline) are also removed through cell death
(Hidalgo et al., 2001
;
Sonnenfeld and Jacobs, 1995b
).
Electron microscopic studies by Sonnenfeld and Jacobs demonstrated that the
majority of cell corpses are expelled from the CNS and engulfed by macrophages
(Sonnenfeld and Jacobs,
1995b
). Cell corpses can also be detected in glial cells both
within and at the surface of the CNS, indicating that glial cells also
contribute to removal of dead cells. These authors also examined
macrophage-less embryos derived from Bic-D mothers and found an
increase in the number of unengulfed cells within the CNS as well as increased
numbers in subperineurial glial cells at the periphery of the CNS. However,
embryos derived from Bic-D mutant mothers have widespread patterning
defects (duplication of posterior structures at the expense of anterior
structures), complicating analysis of the consequences of macrophage loss on
development.
Pvr encodes the Drosophila member of the vertebrate
PDGF/VEGF receptor tyrosine kinase family
(Duchek et al., 2001;
Heino et al., 2001
).
Pvr was initially shown by Duchek et al. to regulate border cell
migration during oogenesis (Duchek et al.,
2001
) and subsequently shown by Cho et al. to control hemocyte
migration (Cho et al., 2002
).
Cho et al. found that hemocytes fail to disperse normally in Pvr
mutant animals and that expression of the Pvr ligand Pvf2 in an ectopic
location can attract hemocytes to the site of Pvf2 expression
(Cho et al., 2002
). We
independently isolated mutations in Pvr and have used our analysis of
Pvr as a starting point to investigate the previously unexplored
function of hemocytes in CNS development.
In this work, we use a combination of gene targeting by homologous recombination and chemical mutagenesis to create mutations in Pvr. We find that Pvr mutations, which disrupt hemocyte migration, cause defects in the patterning of the CNS axon scaffold and the positioning of CNS glia without affecting the pattern of midline glial cell death. We further find that serpent (srp) mutant animals (which lack hemocytes) and animals in which the function of the macrophage scavenger receptor Croquemort has been inhibited both show defects in CNS patterning similar to those of Pvr mutants. Taken together, these data suggest that macrophage-mediated engulfment is necessary for proper Drosophila CNS development.
![]() |
MATERIALS AND METHODS |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Genomic DNA was prepared for long-range PCR by crushing single flies (Canton-S, Pvr knock-out donor, or heterozygous Pvr knock-out) in 50 µl squishing buffer (10 mM Tris-HCl pH 8.2, 1 mM EDTA, 25 mM NaCl, 200 µg/ml Proteinase K). Crushed flies were incubated at 37°C for 30 minutes and Proteinase K was inactivated at 95°C for 2 minutes. Long-range PCR was performed using the Expand Long Template PCR System (Roche). Primers used to confirm homologous recombination were 5'-AATCGTACCGTTGCGAATAAGTGGG-3' (Primer A), 5'-AGAAGCGAGAGGAGTTTTGGCACAGC-3' (Primer B), 5'-TTTGTTCGACGACCTTGGAGCGAC-3' (Primer C) and 5'-TGGATAAAGTTCCATCACCACCACGG-3' (Primer D). Control PCR was performed with primers against Pvr genomic DNA, 5'-CAGTGCAACGCTAAGTGAGCC-3' and 5'-TCTTCACGCAATAGTAGGCTGCC-3'.
The following fly stocks were kindly provided by those indicated: slit(1.0)-lacZ (I. Rebay), Sim-Gal4 (C. Goodman), Gcm-Gal4 (U. Tepass), UAS-PvrDN (P. Rorth), srpneo45 (U. Banerjee) and Df(3L)H99 (Bloomington Stock Center). Homozygous Pvr and srpneo45 mutant embryos were identified through the use of balancers marked with GFP (Bloomington Stock Center).
Southern blotting
Genomic DNA isolated from 25 Canton-S or PvrKO/CyO
flies was digested with PvuII. Digested DNAs were electrophoresed on
an 0.8% agarose gel and transferred by downward capillary transfer to a
Zeta-Probe GT membrane (BioRad). Membrane was subsequently treated following
manufacturer's protocol. The probe template was amplified by PCR using genomic
DNA and primers 5'-ATCGCTCGTATGCCCTACAACG-3' and
5'-CTTCCTGTCAACAATCGCACATTC-3', which span 776 bases of the Pvr
locus. 32P-labelled probe was made from this template using the
DECAprime II Kit (Ambion).
Immunohistochemistry and western blotting
All embryos were stained as described
(Patel, 1994) using: rat
polyclonal antiserum against Repo (Campbell
et al., 1994
) (1:500); mouse monoclonal antibody against
lacZ (1:100; Promega); mouse polyclonal antiserum against Peroxidasin
(Nelson et al., 1994
) (1:500)
and rabbit polyclonal antiserum against Croquemort
(Franc et al., 1999
) (1:500).
The mouse monoclonal antibodies BP102 and 1D4, developed by C. Goodman, were
obtained from the Developmental Studies Hybridoma Bank developed under the
auspices of the NICHD and maintained by The University of Iowa, Department of
Biological Sciences, Iowa City, IA 52242. Secondary antibodies labeled with
HRP, FITC or Cy3 were obtained from Jackson Laboratories. Polyclonal antiserum
against Pvr was produced in rats (Covance) against a 30 kDa peptide containing
the 275 C-terminal amino acids of Pvr fused to a 6XHis tag. Anti-Pvr antiserum
was used at 1:300 on embryos. Fluorescent images were obtained using a Nikon
PCM2000 confocal microscope.
For each lane of western blots, 10 embryos of each genotype were homogenized in phosphate-buffered saline (PBS) (130 mM NaCl, 175 mM Na2HPO4, 60 mM NaH2PO4). Lysates were run on a 9% polyacrylamide gel and transferred to Hybond-P membrane (Amersham Pharmacia). Membranes were blocked in 5% nonfat milk and probed with anti-Pvr antisera diluted 1:2000 and HRP-conjugated goat anti-rat antibody diluted at 1:5000. Blots were stripped and reprobed with rat monoclonal antibody against Elav to confirm that each Pvr mutant lane contained similar or greater amounts of total protein than wild-type control.
RNAi
Inhibition of Crq by RNAi was performed as described
(Kennerdell and Carthew,
1998). Bases 681-1203 and 1209-1737 of Crq cDNA RE02070 (ResGen;
GenBank Accession Number AY070904) were PCR-amplified using primer
5'-GAATTAATACGACTCACTATAGGGAGAGGGACTGATGCCTATGAAAGCTG-3' with
5'-GAATTAATACGACTCACTATAGGGAGAAGCCATCGTAAGTCAGCGACTC-3' and
5'-GAATTAATACGACTCACTATAGGGAGAACTATTCACACGGGCACTGACG-3' with
5'-GAATTAATACGACTCACTATAGGGAGATAATCTGGATGCGTCCATGCAC-3'. The
5'-end of each oligonucleotide contains a T7 RNA polymerase promoter
sequence. dsRNAs were synthesized with T7 RNA polymerase from the MEGAscript
High Yield Transcription Kit (Ambion). dsRNA was injected into Canton-S
embryos at 1 µg/µl. Both Crq dsRNAs gave similar results. Control
embryos injected with dsRNA from the Gal4 gene or Ptp99A gave no detectable
phenotype.
![]() |
RESULTS |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
|
|
Pvr is required for proper CNS axon scaffold formation and glial positioning
We next examined CNS patterning in Pvr mutants. CNS axons in the
Drosophila embryo establish a precise pattern reiterated in each
segment (Fig. 3A). CNS axons
establish two longitudinal tracts that run the length of the embryo on either
side of the midline, with a subset of these axons crossing the midline of the
embryo, forming two commissural axon bundles per segment. CNS axons are guided
in part by signals from glia precisely positioned at the midline and along the
longitudinal tracts (Hidalgo and Booth,
2000a; Kidd et al.,
1999
). Although CNS axon architecture was grossly normal in
Pvr mutants, the precise ladder-like axon scaffold seen in wild-type
embryos was disrupted (Fig.
3B). The scaffold in each segment had a rounded appearance, owing
to changes in the separation between the anterior and posterior commissures
and the longitudinal tracts.
|
|
As Pvr mutants showed disruptions in CNS axon tract shape and
glial cell positioning near the CNS midline, the pathfinding of CNS axons near
the midline in Pvr mutants was examined in greater detail. The
monoclonal antibody 1D4 (mAb 1D4) recognizes the Fasciclin 2 protein
(Van Vactor et al., 1993) and
labels a subset of longitudinal bundles that grow adjacent to the CNS midline.
Mab1D4 is a commonly used tool for assessing axon fasciculation patterns and
detecting inappropriate axon crossing of the CNS midline
(Hidalgo and Brand, 1997
;
Kidd et al., 1998
;
Lin et al., 1994
). Despite the
changes in CNS axon scaffold shape and longitudinal glial distribution
observed in Pvr mutants, no inappropriate axon crossing of the
midline was detected (Fig.
3E,F). In addition, in wild-type animals three major tracts of
Fasciclin 2-positive axons are observed near the dorsal surface of the CNS on
either side of the midline (Fig.
3E). Three major tracts of Fasciclin 2-positive axons were also
observed in Pvr mutants (Fig.
3F).
Although these tracts were relatively normal, they did show very mild
defasciculation in some segments, with axons in Pvr mutants
occasionally separating from one another by greater distances than normal
(Fig. 3F). As disruptions in
longitudinal glial cell development disrupt the formation of these axon
bundles (Hidalgo and Booth,
2000b), the minor axon tract defects observed in Pvr
mutants were not unexpected given the glial cell mispositioning seen in
Pvr mutants (Fig.
3D).
Pvr functions in hemocytes
To investigate the source of the CNS defects in Pvr mutants, we
identified the cell populations expressing Pvr. Pvr protein was detected on
several cell populations during embryonic development. In stage 16 embryos,
Pvr was prominently expressed by cells at the surface of the embryo, as well
as by cells scattered throughout the embryo and by cells at the CNS midline
(Fig. 4A). The large number of
Pvr-expressing cells scattered throughout the embryo were hemocytes, as they
co-expressed the hemocyte marker Peroxidasin
(Fig. 4B-D). The Pvr-expressing
cells at the CNS midline were midline glia (a population distinct from the
Repo-positive glia mentioned above) and were intimately associated with the
CNS commissures (Fig. 4E,F).
Pvr expression could not be detected in PvrKO2,
Pvr5363 or Pvr9742 embryos, confirming
the specificity of the antiserum (Fig.
4G).
|
|
Macrophage function is required for CNS patterning
To further examine the potential contribution of hemocytes to CNS
development, we examined animals mutant for serpent (srp),
which encodes a GATA-family transcription factor required for hemocyte
development (Rehorn et al.,
1996). srpneo45 is a hemocyte-specific allele
of serpent, and srpneo45 animals lack all
hemocytes (Lebestky et al.,
2000
; Rehorn et al.,
1996
). Examination of srpneo45 embryos
demonstrated that not only did srpneo45 mutants lack
macrophages, they also exhibited CNS axon scaffold defects similar to those in
Pvr mutants, with characteristic rounding of commissures
(Fig. 6C). Quantitative
representation of CNS axon tract morphology in srpneo45
mutants confirmed this observation (Table
1). The ratio of the distance between the longitudinal axon tracts
and the distance between the commissural axon tracts in
srpneo45 mutants was significantly different from wild
type (P<0.01), but not significantly different from Pvr
homozygotes (P>0.2) (Table
1). srpneo45 animals also showed longitudinal
glia positioning defects similar to those seen in Pvr mutants
(Fig. 6G). Thus, mutants that
disrupt either hemocyte production or migration cause similar alterations in
CNS morphogenesis.
|
![]() |
DISCUSSION |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
There are several possible explanations for the observed contribution of
macrophages to CNS morphogenesis. As engulfment is capable of promoting cell
death in some situations (Reddien et al.,
2001), the inhibition of macrophage function could potentially
change patterns of cell death. However, previous work found that substantial
cell death still occurs in Drosophila embryos in the absence of
macrophages (Tepass et al.,
1994
). Similarly, we find no alteration in the number of midline
glia in Pvr mutants, suggesting that the death of midline glia
proceeds normally. Acridine Orange staining of developing embryos likewise
shows no detectable alteration in the pattern of dead cell generation (H.C.S.,
C.J.K. and P.A.G., unpublished). Thus, although subtle changes in pattern of
cell death would escape detection by these methods, there is no large-scale
alteration in cell death in Drosophila in the absence of macrophages.
Interestingly, when cell death is blocked in homozygous Df(3L)H99
animals, which lack the cell death promoting genes hid, grim and
reaper, the CNS axon scaffolds are wider than normal
(Zhou et al., 1995
) (H.C.S.,
C.J.K. and P.A.G., unpublished). The phenotypic contrast between the absence
of cell death and the absence of macrophages suggests that macrophages are not
simply required to remove material from the developing midline.
An alternative explanation for the need for macrophage-mediated engulfment
is that the accumulation of cell corpses within the CNS disrupts axon and
glial positioning. Cell corpses could exert a toxic effect or could disrupt
the function of particular cell populations by abnormally accumulating within
these cells. Such possibilities are consistent with Sonnenfeld and Jacobs'
observation that in embryos from Bic-D mutant mothers (which, in
addition to severe embryonic patterning defects, lack macrophages) cell
corpses accumulate at the CNS periphery and in CNS glial cells
(Sonnenfeld and Jacobs,
1995b). In the case of glial cells, it is interesting to note that
the CNS defects of Pvr, serpentneo45 and crq RNAi
animals resemble those of repo mutant animals in which glial
positioning and survival is disrupted
(Campbell et al., 1994
;
Halter et al., 1995
). Thus, a
disruption in glial positioning could lead to the disruption of the CNS axon
scaffold observed in the absence of macrophage-mediated engulfment.
Another possible requirement for the engulfment of dead cells by
macrophages could be that engulfment stimulates the release by macrophages of
factors required for proper CNS morphogenesis. Drosophila macrophages
produce extracellular matrix components such as collagen IV, laminin, papilin,
glutactin and macrophage-derived proteoglycan-1 (MDP-1)
(Fessler et al., 1994;
Gullberg et al., 1994
;
Hortsch et al., 1998
), and the
presence of cell corpses is known to enhance production of at least one of
these, MDP-1 (Hortsch et al.,
1998
). That Pvr, serpentneo45 and crq
RNAi animals all show a mild elongation of the nerve cord (H.C.S., C.J.K. and
P.A.G., unpublished) could reflect defects in extracellular matrix
production.
Cell death is a major component of many morphogenetic processes during
development in vertebrates and invertebrates. However, an essential role for
macrophages in these morphogenetic events has been established only in the
mouse retina, where macrophages appear to act by triggering cell death
(Lang et al., 1994;
Lang and Bishop, 1993
). Unlike
the mouse retina, cell death in the Drosophila CNS does not require
macrophages (Tepass et al.,
1994
). Our data support a different role for macrophages in
Drosophila CNS morphogenesis: mediating clearance of dead cells.
These observations indicate that both cell death and the interaction of
macrophages with cell corpses are required for proper Drosophila CNS
development.
![]() |
ACKNOWLEDGMENTS |
---|
![]() |
REFERENCES |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Abrams, J. M. (2002). Competition and compensation: coupled to death in development and cancer. Cell 110,403 -406.[Medline]
Baehrecke, E. H. (2002). How death shapes life during development. Nat. Rev. Mol. Cell Biol. 3, 779-787.[CrossRef][Medline]
Bangs, P. and White, K. (2000). Regulation and execution of apoptosis during Drosophila development. Dev. Dyn. 218,68 -79.[CrossRef][Medline]
Bello, B. C., Hirth, F. and Gould, A. P. (2003). A pulse of the Drosophila Hox protein abdominal-A schedules the Endo of neural proliferatin via neuroblast apoptosis. Neuron 37,209 -219.[Medline]
Bergmann, A., Tugentman, M., Shilo, B. Z. and Steller, H. (2002). Regulation of cell number by MAPK-dependent control of apoptosis: a mechanism for trophic survival signaling. Dev. Cell 2,159 -170.[Medline]
Campbell, G., Goring, H., Lin, T., Spana, E., Andersson, S.,
Doe, C. Q. and Tomlinson, A. (1994). RK2, a glial-specific
homeodomain protein required for embryonic nerve cord condensation and
viability in Drosophila. Development
120,2957
-2966.
Cho, N. K., Keyes, L., Johnson, E., Heller, J., Ryner, L., Karim, F. and Krasnow, M. A. (2002). Developmental control of blood cell migration by the Drosophila VEGF pathway. Cell 108,865 -876.[Medline]
Duchek, P. and Rorth, P. (2001). Guidance of
cell migration by EGF receptor signaling during drosophila oogenesis.
Science 291,131
-133.
Duchek, P., Somogyi, K., Jekely, G., Beccari, S. and Rorth, P. (2001). Guidance of cell migration by the Drosophila PDGF/VEGF receptor. Cell 107, 17-26.[Medline]
Fadok, V. A. and Chimini, G. (2001). The phagocytosis of apoptotic cells. Semin. Immunol. 13,365 -372.[CrossRef][Medline]
Fessler, L. I., Nelson, R. E. and Fessler, J. H. (1994). Drosophila extracellular matrix. Methods Enzymol. 245,271 -294.[Medline]
Fossett, N., Tevosian, S. G., Gajewski, K., Zhang, Q., Orkin, S.
H. and Schulz, R. A. (2001). The friend of GATA proteins
U-shaped, FOG-1, and FOG-2 function as negative regulators of blood, heart,
and eye development in Drosophila. Proc. Natl. Acad. Sci.
USA 98,7342
-7347.
Franc, N. C. (2002). Phagocytosis of apoptotic cells in mammals, caenorhabditis elegans and Drosophila melanogaster: molecular mechanisms and physiological consequences. Front Biosci. 7,D1298 -1313.[Medline]
Franc, N. C., Dimarcq, J. L., Lagueux, M., Hoffmann, J. and Ezekowitz, R. A. (1996). Croquemort, a novel Drosophila hemocyte/macrophage receptor that recognizes apoptotic cells. Immunity 4,431 -443.[Medline]
Franc, N. C., Heitzler, P., Ezekowitz, R. A. and White, K.
(1999). Requirement for croquemort in phagocytosis of apoptotic
cells in Drosophila. Science
284,1991
-1994.
Gullberg, D., Fessler, L. I. and Fessler, J. H. (1994). Differentiation, extracellular matrix synthesis, and integrin assembly by Drosophila embryo cells cultured on vitronectin and laminin substrates. Dev. Dyn. 199,116 -128.[Medline]
Halter, D. A., Urban, J., Rickert, C., Ner, S. S., Ito, K.,
Travers, A. A. and Technau, G. M. (1995). The homeobox gene
repo is required for the differentiation and maintenance of glia function in
the embryonic nervous system of Drosophila melanogaster.
Development 121,317
-332.
Hanks, S. K., Quinn, A. M. and Hunter, T. (1988). The protein kinase family: conserved features and deduced phylogeny of the catalytic domains. Science 241, 42-52.[Medline]
Heino, T. I., Karpanen, T., Wahlstrom, G., Pulkkinen, M., Eriksson, U., Alitalo, K. and Roos, C. (2001). The Drosophila VEGF receptor homolog is expressed in hemocytes. Mech. Dev. 109,69 -77.[CrossRef][Medline]
Hidalgo, A. and Booth, G. E. (2000a). Glia
dictate pioneer axon trajectories in the Drosophila embryonic CNS.
Development 127,393
-402.
Hidalgo, A. and Booth, G. E. (2000b). Glia
dictate pioneer axon trajectories in the Drosophila embryonic CNS.
Development 127,393
-402.
Hidalgo, A. and Brand, A. H. (1997). Targeted
neuronal ablation: the role of pioneer neurons in guidance and fasciculation
in the CNS of Drosophila. Development
124,3253
-3262.
Hidalgo, A., Kinrade, E. F. and Georgiou, M. (2001). The Drosophila neuregulin vein maintains glial survival during axon guidance in the CNS. Dev. Cell 1, 679-690.[Medline]
Hopkinson-Woolley, J., Hughes, D., Gordon, S. and Martin, P.
(1994). Macrophage recruitment during limb development and wound
healing in the embryonic and foetal mouse. J. Cell
Sci. 107,1159
-1167.
Hortsch, M., Olson, A., Fishman, S., Soneral, S. N., Marikar, Y., Dong, R. and Jacobs, J. R. (1998). The expression of MDP-1, a component of Drosophila embryonic basement membranes, is modulated by apoptotic cell death. Int. J. Dev. Biol. 42, 33-42.[Medline]
Huang, Z., Shilo, B. Z. and Kunes, S. (1998). A retinal axon fascicle uses spitz, an EGF receptor ligand, to construct a synaptic cartridge in the brain of Drosophila. Cell 95,693 -703.[Medline]
Hume, D. A., Perry, V. H. and Gordon, S. (1983). Immunohistochemical localization of a macrophage-specific antigen in developing mouse retina: phagocytosis of dying neurons and differentiation of microglial cells to form a regular array in the plexiform layers. J. Cell Biol. 97,253 -257.[Abstract]
Jacobs, J. R. (2000). The midline glia of Drosophila: a molecular genetic model for the developmental functions of glia. Prog. Neurobiol. 62,475 -508.[CrossRef][Medline]
Jacobs, J. R. and Goodman, C. S. (1989). Embryonic development of axon pathways in the Drosophila CNS. I. A glial scaffold appears before the first growth cones. J. Neurosci. 9,2402 -2411.[Abstract]
Jacobson, M. D., Weil, M. and Raff, M. C. (1997). Programmed cell death in animal development. Cell 88,347 -354.[Medline]
Jiang, C., Baehrecke, E. H. and Thummel, C. S.
(1997). Steroid regulated programmed cell death during Drosophila
metamorphosis. Development
124,4673
-4683.
Johnson, L. N., Noble, M. E. and Owen, D. J. (1996). Active and inactive protein kinases: structural basis for regulation. Cell 85,149 -158.[Medline]
Kennerdell, J. R. and Carthew, R. W. (1998). Use of dsRNA-mediated genetic interference to demonstrate that frizzled and frizzled 2 act in the wingless pathway. Cell 95,1017 -1026.[Medline]
Kidd, T., Bland, K. S. and Goodman, C. S. (1999). Slit is the midline repellent for the robo receptor in Drosophila. Cell 96,785 -794.[Medline]
Kidd, T., Brose, K., Mitchell, K. J., Fetter, R. D., Tessier-Lavigne, M., Goodman, C. S. and Tear, G. (1998). Roundabout controls axon crossing of the CNS midline and defines a novel subfamily of evolutionarily conserved guidance receptors. Cell 92,205 -215.[Medline]
Klambt, C., Jacobs, J. R. and Goodman, C. S. (1991). The midline of the Drosophila central nervous system: a model for the genetic analysis of cell fate, cell migration, and growth cone guidance. Cell 64,801 -815.[Medline]
Lang, R., Lustig, M., Francois, F., Sellinger, M. and Plesken,
H. (1994). Apoptosis during macrophage-dependent ocular
tissue remodelling. Development
120,3395
-3403.
Lang, R. A. and Bishop, J. M. (1993). Macrophages are required for cell death and tissue remodeling in the developing mouse eye. Cell 74,453 -462.[Medline]
Lebestky, T., Chang, T., Hartenstein, V. and Banerjee, U.
(2000). Specification of Drosophila hematopoietic lineage by
conserved transcription factors. Science
288,146
-149.
Lin, D. M., Fetter, R. D., Kopczynski, C., Grenningloh, G. and Goodman, C. S. (1994). Genetic analysis of Fasciclin II in Drosophila: defasciculation, refasciculation, and altered fasciculation. Neuron 13,1055 -1069.[Medline]
Lohmann, I., McGinnis, N., Bodmer, M. and McGinnis, W. (2002). The Drosophila Hox gene deformed sculpts head morphology via direct regulation of the apoptosis activator reaper. Cell 110,457 -466.[Medline]
Morris, L., Graham, C. F. and Gordon, S. (1991). Macrophages in haemopoietic and other tissues of the developing mouse detected by the monoclonal antibody F4/80. Development 112,517 -526.[Abstract]
Nambu, J. R., Lewis, J. O., Wharton, K. A., Jr and Crews, S. T. (1991). The Drosophila single-minded gene encodes a helix-loop-helix protein that acts as a master regulator of CNS midline development. Cell 67,1157 -1167.[Medline]
Nelson, R. E., Fessler, L. I., Takagi, Y., Blumberg, B., Keene, D. R., Olson, P. F., Parker, C. G. and Fessler, J. H. (1994). Peroxidasin: a novel enzyme-matrix protein of Drosophila development. EMBO J. 13,3438 -3447.[Abstract]
Noordermeer, J. N., Kopczynski, C. C., Fetter, R. D., Bland, K. S., Chen, W. Y. and Goodman, C. S. (1998). Wrapper, a novel member of the Ig superfamily, is expressed by midline glia and is required for them to ensheath commissural axons in Drosphila. Neuron 21,991 -1001.[Medline]
Patel, N. H. (1994). Imaging neuronal subsets and other cell types in whole-mount Drosophila embryos and larvae using antibody probes. In Methods in Cell Biology, Vol. 44 (ed. E. A. Fyrberg), pp.445 -487. San Diego: Academic Press.[Medline]
Reddien, P. W., Cameron, S. and Horvitz, H. R. (2001). Phagocytosis promotes programmed cell death in C. elegans. Nature 412,198 -202.[CrossRef][Medline]
Rehorn, K. P., Thelen, H., Michelson, A. M. and Reuter, R.
(1996). A molecular aspect of hematopoiesis and endoderm
development common to vertebrates and Drosophila.
Development 122,4023
-4031.
Rong, Y. S. and Golic, K. G. (2000). Gene
targeting by homologous recombination in Drosophila.
Science 288,2013
-2018.
Rong, Y. S. and Golic, K. G. (2001). A targeted
gene knockout in drosophila. Genetics
157,1307
-1312.
Sonnenfeld, M. J. and Jacobs, J. R. (1995a).
Apoptosis of the midline glia during Drosophila embryogenesis: a correlation
with axon contact. Development
121,569
-578.
Sonnenfeld, M. J. and Jacobs, J. R. (1995b). Macrophages and glia participate in the removal of apoptotic neurons from the Drosophila embryonic nervous system. J. Comp. Neurol. 359,644 -652.[Medline]
Tepass, U., Fessler, L. I., Aziz, A. and Hartenstein, V.
(1994). Embryonic origin of hemocytes and their relationship to
cell death in Drosophila. Development
120,1829
-1837.
Van Vactor, D. V., Sink, H., Fambrough, D., Tsoo, R. and Goodman, C. S. (1993). Genes that control neuromuscular specificity in Drosophila. Cell 73,1137 -1153.[Medline]
Wood, W., Turmaine, M., Weber, R., Camp, V., Maki, R. A.,
McKercher, S. R. and Martin, P. (2000). Mesenchymal cells
engulf and clear apoptotic footplate cells in macrophageless PU.1 null mouse
embryos. Development
127,5245
-5252.
Zhou, L., Hashimi, H., Schwartz, L. M. and Nambu, J. R. (1995). Programmed cell death in the Drosophila central nervous system midline. Curr. Biol. 5, 784-790.[Medline]
Related articles in Development: