In This IssueAuthor(s): Nicole LeBrasseurSource: The Journal of Cell Biology, Vol. 159, No. 1 (Oct. 14, 2002), pp. 12-13Published by: The Rockefeller University PressStable URL: http://www.jstor.org/stable/1621235 .

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Neutrophils are gluttonous little cells-they will engulf just about any small particle that comes their way. But eating

up an entire bacterium takes significant effort. On page 181, Dewitt and Hallett report that neutrophils need integrins and a Ca^ boost to build a mouth that is up to the task.

' r Inteerin (yellow) freed by calpain (left) ment components, aggregates annind the closing phagocytic cup. which are the or o / r

targets for integrin binding/ the phagocytic cups close around the bugs more rapidly. The authors found that this speediness

correlated with a sequence of Ca24^ changes during both cup formation and closure. At cup formation, complement binding to integrin initiated the release of local Ca^ stores near the contact site. Local increase caused an unknown PI3K-

dependent signal to launch a cell-wide influx of Ca^ just before particle enclosure. Although Hallett speculates that a

locally activated phospholipid could stimulate Ca^ channels as it diffuses around the plasma membrane, he admits that diffusion of the P13K product, PIP3, is probably too slow to account for the rapid Ca^ influx.

Global Ca^ changes effected the rapid engulfment of

tagged particles by releasing integrin receptors throughout the cell membrane. These receptors were then free to diffuse to the phagocytic site, where they could help hasten

cup closure. Since calpain inhibitors blocked integrin diffusion and rapid phagocytosis, the authors conclude that Ca2'1' may stimulate calpain protease activity throughout the cell. Calpain could cleave talin or some other, perhaps neutrophil-specific, protein that links integrin to the actin

cytoskeleton. *

' I ^ he need for Ran and GTP

JL in nuclear import reactions JH^II

depends on the size of the ^^Eili

imported cargo, according to JBF^ new results by Lyman et al.

(page 55), who show that

large proteins in particular ^.fflBM^I

are susceptible to stalling

mid-import when the small

GTPase Ran is missing. Si^| Imported proteins, in associ- ^^Bl

ation with cargo receptors such

as importin P or transportin, w?^

pass from one nucleoporin ro

protein to another within the nuclear

pore complex (NPC) on their way to the

nucleoplasm. Ran was already known

to help cargo detach from its receptor in

the nucleus. The new results give Ran a

second unexpected function within the

center of the channel.

This function stems from the ability of

RanGTP to bind to cargo receptors. When

Ran or GTP was absent from importin a/P and transportin import reactions, or

^ : receptor before the cargo also

*"iir ̂ ^- ^s ^'^^^d leave the receptor/ - ^ :' ^ cargo complex free to bind to

^ i^ another nucleoporin.

l^i ^ | Receptor/cargo movement

^^j^^ from one nucleoporin to the

next within the NPC is thought

I to occur by passive diffusion.

Increasing the off-rate of

receptors from nucleoporins

through RanGTP binding would be especially favorable

for the import of large cargoes ( o om). because they need more off-time

to diffuse within the restricted space of the

NPC to the next nucleoporin. The model

also predicts that receptors with low

affinities for nucleoporins will not rely on

Ran for importing larger proteins, which

may explain why other studies have shown

that import via snurportin does not require Ran or GTP. Now, biophysical studies are

needed to confirm that RanGTP favors

dissociation of receptor from nucleoporin rather than receptor from cargo. *

irgo is not imported into the nucleus without GTP

if the receptors were unable to bind to

Ran, large cargoes got stuck on the

cytoplasmic side of the NPC. Previous

in vitro studies have shown that RanGTP

binding to cargo receptors dissociates

receptors from both cargo and nucleoporins. With their new results, the authors

propose that RanGTP may also bind to

receptors in a way that selectively dissociates

nucleoporins but not cargo. Rapid

subsequent dissociation of Ran from the

12 The Journal of Cell Biology | Volume 159, Number 1, 2002

In This Issue

Calcium increases the appetite

Big proteins Ran into import difficulties

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Neutrophils choose the right direction Two distinct signaling pathways operate sequentially, thus guiding neutrophils to bacterial

invaders, according to Heit et al. on page 91. Although one pathway lures neutrophils in the right general direction, the other pathway dominates when the prey is within reach.

Neutrophils leave the bloodstream 40 n 40 - and migrate to infection sites by IL 8 Induced Aid Phosphorylation Induced ? J

35-^ Aktphosphoryiation 35 byiL-8+iMLp following intermediary chemokine

? 30 4 g3o signals (such as IL-8) that are

*j.25 ̂ |,25 generated by damaged host cells.

1.20 \ I. 20- End-target signals like fMLP or

115 - \ I 15 -- LPS, either released by the pathogen ^ 10 k-^^^ ^ io\ itself or by the body in response to

5 A L ^^^^^^-^s. 5'?* *? Ae bacteria, also attract neutrophils.

o f ,,,,,, o Since the blood cells prefer to pur- 0 10 20 30 40 50 60 0 10 20 30 40 50 60 j 1 1 1 * 1

Tune (nunutes) Time (nunutes) sue CIld-targCtS when both Signals

End targets win the migration battle by blocking Akt are

P^ ;the authors speculated,

phosphorylation (right), correcdy, that different pathways controlled the two responses.

The pathways can be distinguished by their kinase-dependence. Migration to intermediary chemoattractants depended on PI3K, which phosphorylated Akt. End-targets induced

migration via p38 MAPK and inhibited Akt phosphorylation, even at low concentrations.

The result is that neutrophils ignore intermediate molecules when bacteria are in the vicinity. The authors speculate that migration may change depending on the a integrin subunit

recruited-the CD1 la recruited by PI3K may be subservient in migration competence to

the CD1 Ib recruited by p38 MAPK. The findings could explain why the leakage of bacterial products into the bloodstream can cause such severe problems, ^ ^ as these chemicals keep neutrophils from following i _ _

intermediary signals into the surrounding tissue. * j | ^H

A GAP in COPI vesicle 11 1^ * ^ - - ^ - * * F* 11 I ARFGAP1 + - + -

formation is filled ^p GTp,s The effect of ARF1 GTPase-activating protein (ARFGAP1) Without ARFGAP1 or GTP,

on vesicle formation is making a turn-around. Contrary vesicles cannot load their cargo. to previous theories, GAP does not antagonize COPI coat recruitment. As shown by Yang et al. on page 69, its newly discovered function reveals conservation among GAPs in anterograde and retrograde transport pathways.

COPI vesicle formation is initiated by the ADP-Ribosylation Factor (ARF) family of small

GTPases, which recruits COPI coatomer subunits to Golgi membranes. ARFGAP1 inactivates ARF1 by stimulating GTP hydrolysis. Because COPI vesicles formed in vitro using nonhydrolyzable GTP (GTP-yS) or GTP-bound ARF1 do not uncoat, it has been inferred that ARFGAP1 stimulates uncoating, and thus inhibits vesicle formation. But the new

experiments, using hydrolyzable GTP, provide a clearer view of the ARFGAP1 function.

Yang et al. found that ARFGAP1 had the opposite effect of what was previously thought-it stimulated vesicle formation. ARFGAP1 was required for cargo sorting and was found on vesicles at levels exceeding even that of COPI. Thus, ARFGAP1 is actually a COPI coat component, similar to the GAP Sec23p on COPII vesicles. In the COPI case, GTP was required for ARF1 to bring ARFGAP1 to the site of vesicle formation. GTP-yS blocked this recruitment, and thus blocked vesicle formation altogether. Additional

shearing manipulations in previous in vitro experiments may have masked the requirement for ARFGAP1 in vesicle formation by releasing abnormal vesicles. *

TEXT BY NICOLE LEBRASSEUR

LEBRASN@ROCKEFELLER.EDU

Everyone needs a home

n apoptosis-promoting factor leads cells to suicide

by making them feel homeless.

With their report on page 169,

Wang et al. finally explain how a

long-known death kinase initiates

apoptosis. 3 - The kinase in

question is

^ 9 death associ- 0 -^-. .^ ~"~j ated protein 1 -I "1 ^ kinase

^ * | (DAPK), a

0 I I P051^ , vector DAPK regulator or

Integrin activation apoptosis

(black bars) blocks induced by DAPK-induced many stimuli, apoptosis. including c-myc and TGF-(B. Although DAPK was cloned several years

ago, its function in cell death has

remained elusive. The new results reveal that DAPK has the unusual

ability to initiate anoikis, a form

of apoptosis induced in unattached

cells, by interfering with integrin

signaling. Not one to dally with down-

stream signals, DAPK starts at the

top-by altering integrin structure.

Wang et al. found that DAPK

locks integrin in an inactive state, thus suppressing epithelial cell

adhesion to the extracellular matrix

(ECM). As a result, the ECM

survival pathway initiated by

integrin and mediated by FAK

was blocked, resulting in cell death. DAPK effects on adherence and

survival were reversed by activating integrin or by expressing active

FAK. Carcinoma cells, which

are resistant to anoikis, were

unaffected by DAPK. Wang et al.

do not yet know whether DAPK

binds to the cytoplasmic tails of

integrin or how it might inactivate the receptor. *

In This Issue 13

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