Download Trafficking of phosphatidylinositol by phosphatidylinositol transfer

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Biochem. Soc. Symp. 74, 259–271
(Printed in Great Britain)
© 2007 The Biochemical Society
Trafficking of
phosphatidylinositol by
phosphatidylinositol transfer
proteins
Shamshad Cockcroft1
Lipid Signalling Group, Department of Physiology, University College London,
London WC1E 6JJ U.K.
Abstract
PtdIns is synthesized at the endoplasmic reticulum and its intracellular
distribution to other organelles can be facilitated by lipid transfer proteins
[PITPs (phosphatidylinositol transfer proteins)]. In this review, I summarize
the current understanding of how PITPs are regulated by phosphorylation,
how can they dock to membranes to exchange their lipid cargo and how cells
use PITPs in signal transduction and membrane delivery. Mammalian PITPs,
PITPα and PITPβ, are paralogous genes that are 94% similar in sequence.
Their structural design demonstrates that they can sequester PtdIns or
PtdCho (phosphatidylcholine) in their hydrophobic cavity. To deliver the
lipid cargo to a membrane, PITP has to undergo a conformational change at
the membrane interface. PITPs have a higher affinity for PtdIns than PtdCho,
which is explained by hydrogen-bond contacts between the inositol ring of
PtdIns and the side-chains of four amino acid residues, Thr59, Lys61, Glu86
and Asn90, in PITPs. Regardless of species, these residues are conserved in all
known PITPs. PITP transfer activity is regulated by a conserved serine residue
(Ser166) that is phosphorylated by protein kinase C. Ser166 is only accessible for
phosphorylation when a conformational change occurs in PITPs while docking
at the membrane interface during lipid transfer, thereby coupling regulation
of activity with lipid transfer function. Biological roles of PITPs include their
ability to couple phospholipase C signalling to neurite outgrowth, cell division
and stem cell growth.
1
email [email protected].
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S. Cockcroft
Introduction
PITPs (phosphatidylinositol transfer proteins) were first identified as
proteins that were able to bind and transport phospholipids, in particular PtdIns
and PtdCho (phosphatidylcholine). The first mammalian PITP was purified from
bovine brain [1] and its sequence was determined in 1989 [2]. It encoded a protein
of 271 amino acids, which showed no sequence similarity with other known
proteins. Previously, a PITP that facilitates the transfer of PtdIns and PtdCho
in vitro has been isolated from the cytosol of yeast, Saccharomyces cerevisae
[3]. This protein is encoded by the SEC14 gene and is required for transport of
secretory proteins from the Golgi complex in yeast [4]. However, it is noteworthy
that, although mammalian PITPs are similar to yeast Sec14p in lipid binding and
transfer properties, these proteins share no sequence or structural homology.
Functionally, mammalian and yeast PITPs can partially overlap, suggesting that
the common feature of lipid transfer is highly relevant to PITP function [5–7].
Since the discovery of PITPs, their function has been coupled to phospholipid, particularly phosphoinositide, metabolism, linking it to lipid signalling and
membrane traffic [8–10]. Despite a detailed knowledge of the biochemical activities of PITPs in vitro, a molecular understanding of how PITPs fulfil their functions in intact cells and in complex organisms remains poor. The current view is
that it is unlikely that PITPs are passive mediators of phospholipid transfer and,
instead, their activity is harnessed in specific phospholipid metabolic pathways
that can have an impact on both lipid signalling and vesicular delivery [11]. In
mammals, three soluble PITPs and two membrane-anchored proteins that harbour a PITP domain have been identified (Table 1). Deficiencies in specific PITPs
in different organisms are associated with neurodegeneration, abnormalities in
cell division and other specific disorders (Table 2). In the present paper, recent
advances in PITP structural biology are reviewed, highlighting the important
features that are the hallmark of all PITPs. Moreover, the available structures
clarify how PITPs undergo a conformational change when they interact with the
membrane to exchange their lipid cargo. During lipid exchange, residues that are
normally inaccessible to PKC (protein kinase C) become accessible and phosphorylation can then regulate PtdIns transfer activity.
Mammalian PITPs can be categorized into Class I and Class II based
on sequence differences. Class I PITPs contain a single domain and in mammals, two highly related 35 kDa PITPs (α and β) are found (Table 1). PITPβ is
alternatively spliced such that 16 residues at its C-terminus are different in the
two variants. Class II PITPs comprise the members of the RdgB family, the soluble RdgBβ and two RdgBα (I and II) isoforms. RdgBα are large proteins (approx.
160 kDa) where an N-terminal PITP domain is followed by several recognized
domains including a FFAT motif [two phenylalanines (FF) in an Acidic Tract],
which targets the protein to the endoplasmic reticulum by interacting with VAP
(vesicle-associated membrane protein-associated protein) [12], and six stretches
of hydrophobic residues, which are responsible for membrane association. In
multicellular organisms, including Caenorhabditis Elegans (worms), Drosophila
(flies), Xenopus (frog) and Zebrafish, both Class I and II PITPs are found, whereas unicellular organisms appear to have only Class I PITPs. Examples of the
© 2007 The Biochemical Society
PITPNA
PITPNB
PITPNC1
PITPNM1
PITPNM2
17p13.3
22q12
17q24.2
11q13.1
12q24.31
PITPα (270)
PITPβ-sp1 (269)
PITPβ-sp2 (270)
RdgBβ-sp1 (332)
RdgBβ-sp2 (268)
RdgBα1 (1244) (Nir2; PITPnm)
RdgBα2 (1349) (Nir3; PITPnm2)
Gene name
Chromosomal localization
Proteins with a PITP domain in humans (a.a.)
Q00169
P48739
P48739-2
Q9UKF7
Q96I07
O00562
Q9BZ72
Accession No.
100
77
76
39
40
45
43
Identity(%)
Table 1 Proteins (and splice variants) found in the human genome that contain a PITP domain. Accession No. refers to the
TREMBLE or NCBI databases. sp indicates splice variants; a.a., no. of amino acids.
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© 2007 The Biochemical Society
PITP
PITPα
PITPβ
RdgBαI
RdgBαII
RdgBα
PITP
PITP
Organism
Mice
Mice
Mice
Mice
© 2007 The Biochemical Society
Drosophila
Drosophila
Planaria
(flatworm)
PITPα is enriched in the brain;
both reduction in PITPα levels and
its complete knockout cause
neurodegeneration and juvenile death
PITPβ is Golgi localized
The mammalian homologue can rescue
the mutant fly phenotype
Expressed primarily in retina and
dentate gyrus but cannot rescue mutant
fly phenotype
The RdgB mutant fly shows a defective
light response as measured by
electroretinograms and subsequently
suffers a light-dependent photoreceptor
degeneration
Abnormalities in mitotic spindle
formation and in the actomyosin
contractile ring. A single Class I PITP
is found in this organism which cannot
be classified as PITPα or PITPβ.
PITP identified in a RNAi screen
performed to identify genes required for
regeneration following wounding
Comments
[33]
[32]
[61]
[60]
[58]
[59,60]
[56,57]
References
262
Regeneration; stem cell proliferation
Cytokinesis
Retinal degeneration
No obvious phenotype
Cell lethal
Embryonic lethal
Neurodegeneration; intestinal
malabsorption disorders;
glucose homoeostasis
Abnormalities
Table 2 Phenotypes observed upon deletion of specific PITPs
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latter include Giardia Lamblia (Diplomonad), Encephalitozoon (Microsporidia)
[13], red alga Cyanidioschyzon merolae [14] and Dictyostelium Discoideum [15].
In Plasmodium falcipurum (NP_707395.1), Plasmodium Berghei (XP_674861.1)
and Plasmodium Yoelii Yoelii (EAA20463.1), a single PITP domain that is part
of a larger protein and contains a pleckstrin homology domain has been identified in the NCBI database.
Defining the PITP domain
The defining feature of PITPs is the ability to bind to PtdIns and, to a lesser
extent, PtdCho. The structures of the soluble PITPs, PtdCho-bound PITPα
and PITPβ [16,17], PtdIns-bound PITPα [18] and apo-PITPα that is devoid
of its lipid cargo [19], have been solved. The PITP domain consists of eight
β-strands and seven α-helices; the β-sheets form a large concave sheet flanked
by two long α-helices, A and F, that form the lipid-binding cavity (Figure 1A).
Figure 1 (A) Three-dimensional structure of human PITPα bound to PtdIns.
The lipid-binding site comprises of an eight-stranded concave β-sheet (coloured
yellow) flanked by two α-helices, A and F (coloured blue), and sequesters the
PtdIns molecule (space-filled, coloured wheat). The C-terminal G-helix, followed
by 11 amino acids (coloured red), together constitute the lid that keeps the lipid
trapped in the hydrophobic cavity. The Ser166 residue that can be phosphorylated
by PKC is shown in the regulatory loop (coloured green). Two tryptophan residues
(203/204WW) that are thought to make contact with the membrane interface
are shown (coloured gold). Insertion of these two residues in the membrane
may provide the driving force for the ‘lid’ to move away, exposing the lipid to the
membrane. (B) The amino acid side-chains responsible for co-ordinating the inositol
headgroup are indicated, as are the residues important for making contact with
the phosphate group in the phospholipid (dotted green lines). Green=phosphate,
magenta=lipid carbon, red=oxygen, blue=nitrogen and yellow=sulfur. The inositol
ring is numbered 1–6. Single letter amino acid codes are used.
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S. Cockcroft
This cavity is closed by a ‘lid’ composed of a C-terminal α-helix (G-helix) and
an 11 amino acid tail. The headgroup of the bound lipid is buried deep in the
lipid-binding cavity with the acyl chains pointing towards the lid. The lipid
would thus exit out of its binding pocket with its acyl tail inserting into the
membrane first. An important point that emerges from the structure of PITPs
is that the lipid cargo is completely shielded from the aqueous phase and that its
headgroup is not accessible for phosphorylation by lipid kinases. The structures
of the lipid-loaded PITPs (α and β) are very similar [16], suggesting that the
mechanisms of lipid loading and release are very similar for these PITPs.
The PtdIns- and PtdCho-loaded structures of PITPs provide the framework
for understanding why the affinity of PITPs for PtdIns is much greater than that
for PtdCho [20,21]. Examination of the PITPα–PtdIns structure identified sidechains within the lipid-binding pocket that interact via a specific set of hydrogen
bonds only with the inositol ring of PtdIns (Figure 1B). Amino acid residues that
are important for inositol binding are Thr59, Lys61, Glu86 and Asn90. Mutations
made at each of these residues to prevent the hydrogen bond interactions
between the inositol ring of PtdIns and PITPα confirm the importance of these
residues for PtdIns, but not PtdCho, binding and transfer. These residues are
absolutely conserved in all 81 PITP sequences that are currently available in the
database. Another conserved feature is the amino acid residues that make contact
with the phosphate group of the phospholipid (Gln22, Thr97, Thr114 and Lys195)
(Figure 1B). Thr97 and Lys195 residues are conserved in all PITP sequences examined,
whereas Gln22 and Thr114 are replaced in a minor number of sequences through a
conservative substitution (threonine with serine and glutamine with lysine).
Interaction of PITPs with membrane surfaces for lipid
exchange
In the cytosol, PITPs spend a long time in the ‘closed’ conformation.
However, to execute their lipid transfer function, PITPs have to interact with
membranes. Previous studies have shown that the C-terminus of PITPs plays a
crucial role in this process [22]. Evidence that PITPα undergoes a conformational
change during binding to membranes comes from the observation that the
C-terminus of PITPα is susceptible to proteolytic cleavage by trypsin only
when PITPα is bound to phospholipid vesicles [23]. At the membrane, the
conformational change in PITPs would result in accessibility of the lipid binding
cavity essential for the release of the phospholipid cargo. The crystal structure
of apo-PITPα provides a partial insight into the events that could unfold at the
membrane. In this lipid-free structure, the G α-helix has swung outwards by
approx. 20°. Additional insights were obtained from the observation that two
PtdIns-loaded PITPα molecules are arranged as an end-to-end dimer in the
crystal structure [18]. The dimer interface is a potential site for membrane
interaction and contains two aromatic tryptophan residues (Trp203 and Trp204)
(see Figure 1A), which are chemically favourable for interaction with a membrane
interface. Mutation of the tryptophan residues to alanine severely impairs
PtdIns and PtdCho transfer without disrupting the lipid binding ability of
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PITPα [18]. Alignment of PITP domains from all known sequences indicates
that, although Trp203 and Trp204 are not conserved, a hydrophobic residue is
almost always present in at least one (usually both) of these positions. I suggest
that, in the closed configuration, PITPs initially interact with the interfacial
region of the membrane through these two adjacent tryptophans. Upon
membrane association, and possibly membrane insertion, the C-terminus of
PITPs moves to expose the hydrophobic lipid-binding cavity to the membrane.
In its membrane-associated conformation, PITP can exchange its lipid cargo.
Phosphorylation sites in the PITP domain and their
functional relevance
Treatment of cells with PMA leads to phosphorylation of PITPα at
Ser166 (Figure 1A). In addition, both recombinant and brain-derived PITPα is
phosphorylated at this residue in vitro, although at a lower stoichiometry [24,25].
Ser166-phosphorylated PITPs can also be isolated from brain cytosol, confirming
that phosphorylation takes place in vivo [25]. The Ser166 residue is located in the
regulatory loop (coloured green in Figure 1A), and the side-chain of this residue
is located in a small pocket formed by amino acid residues 165–172. In the soluble
structure of PITPs, Ser166 is inaccessible for phosphorylation by PKC, explaining
the low phosphorylation stoichiometry. For phosphorylation to occur PITPs
must undergo a conformational change and most probably this occurs at the
membrane interface during lipid exchange (discussed above). In the structure of
the apo-PITPα, Ser166 is also inaccessible suggesting that, at the membrane, the
protein undergoes a more dramatic change than is seen in the apo-structure.
Alignment of the 81 PITPα-related sequences identified in mammals, fish,
amphibians, flies, soil amoebae, red photosynthetic algae and parasites indicates
that Ser166 is conserved in 77 out of 81 sequences. In 73 sequences, a consensus
sequence EDP(X)4S(X)K/R(X)2RG (where X is any amino acid) can be identified.
Ser166 is not conserved in one of the C. elegans and Caenorhabditis briggsae
PITPs and in the PITP from Encephalitozoan cuniculi and Giardia. This strong
conservation indicates that the regulatory loop is important in PITP function
and phosphorylation plays a fundamental role in most PITP-related molecules.
A major function of PITPs is to supply PtdIns, which once phosphorylated
to PtdIns(4,5)P2 is the substrate for PLC (phospholipase C) [8]. Both PITPα and
PITPβ are found at the plasma membrane following activation with epidermal
growth factor, which also activates PLC [11,26]. Thus PITPs would be positioned
close to the site of diacylglycerol generation and thus PKC activation. The effect
of phosphorylation at Ser166 on PITPα is to reduce PtdIns transfer [25]. Mutation
of PITPα Ser166 to either alanine or glutamate also abolishes transfer activity. The
equivalent residue in PITPβ is Ser165 and again mutation of this residue abolishes
lipid transfer activity. Thus similar to PITPα, PITPβ is controlled through
phosphorylation by PKC. Phosphorylation of PITPs would result in a reduction
of substrate supply, providing a mechanism for negative feedback regulation
of PLC activation by PKC. It is well-established that pretreatment with PMA
disrupts receptor-mediated PLC signalling in most cell types and our data
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suggest that phosphorylation of PITPs at Ser166 provides a common regulatory
mechanism regardless of the receptor or the PLC isoform involved [25].
The observation that mutation of Thr59 has profound effects on phosphorylation of Ser166 is indicative of the point in the cycle of lipid exchange at which
PITPs get phosphorylated in vivo. In PITPα–PtdIns, Thr59 forms a hydrogen
bond with the headgroup of PtdIns (Figure 1B), and when it is mutated to
alanine, serine or glutamate, the mutants are still capable of binding to PtdCho,
whereas PtdIns binding is either abolished (T59E) or reduced (T59A and
T59S). In vitro, PITPα-T59A gets phosphorylated by PKC on residue Ser166
to a higher stochiometry compared with wild-type PITPα. More striking is
the degree of phosphorylation observed in permeabilized cells stimulated with
GTP[S] (guanosine 5’-[δ-thio] triphosphate); GTP[S] activates the endogenous
PLCβ to make diacylglycerol that activates the endogenous PKC. Under conditions when PLC is activated, phosphorylation of PITPα-T59A at Ser166 is
greater than for wild-type PITPα. The reason behind these differences between
wild-type PITPα and the mutant, PITPα-T59A lies in the presence of a distinct
structural form that can be separated on native gels [25]. The observation that
changes in the Thr59 residue can have such profound effects on Ser166 accessibility to PKC suggests that the available structures of PITPs are not adequate
to understand the conformational structures that PITPs might undergo. The
available data suggest that when the protein is devoid of its lipid cargo, Ser166 is
accessible for phosphorylation by PKC.
PITPβ can also be phosphorylated at an additional residue, Ser262, which is
substituted by proline in PITPα. Ser262 lies at the end of the G helix and is exposed to the cytosol. Mutation of this residue to alanine has no effect on PtdIns
transfer. In cells, Ser262 is constitutively phosphorylated and can be dephosphorylated upon prolonged treatment with the PKC inhibitor GF 109203X [27,28].
It has been reported that the Golgi localization of PITPβ is dependent on Ser262
phosphorylation [27]. However, the PITPβ splice variant (PITPβ-sp2), which
lacks Ser262, still localizes on the Golgi [28]. Moreover, the S262A mutant is also
Golgi-localized as is the dephosphorylated wild-type PITPβ [28]. The major
difference between PITPα and the two splice variants of PITPβ is at the extreme
C-terminus (coloured red in Figure 1A) that forms the lid to the hydrophobic
cavity, and thus, the Golgi localization of PITPβ variants might be determined by
interactions between this region and the membrane. Whether a Golgi protein or a
specific lipid domain is required for this interaction is not obvious. What is clear,
however, is that PITPβ interactions with the Golgi membrane are dynamic, as
monitored by the FRAP (fluorescence recovery after photobleaching) technique
and, when cells are broken to make membrane preparations, very little PITPβ
remains membrane associated [11].
Biological functions of PITPs
The PITP domain is designed to bind to a molecule of PtdIns, and the most
parsimonious function of PITP is probably to deliver PtdIns to specific membrane
compartments. Target membranes can convert PtdIns by phosphorylation into
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a variety of phosphoinositides including PtdIns(4,5)P2. Since phosphoinositides
play critical roles in many aspects of cell biology, the potential role for PITPs in
these processes is enormous. Previous studies performed in permeabilized cells
indicate that PITPs play roles in PLC signalling [29], exocytosis [30] and vesicle
formation [7]. In these studies, both PITPα and PITPβ were effective. Although
the phenotypes of the PITPα- and PITPβ-knockout mice are distinct (Table 2),
the tissue distribution of these proteins, as well as their cellular localization,
varies; PITPβ localizes to the Golgi compartment where it probably plays a
specific role. It is thus highly possible that, as well as overlapping functions,
PITPα and PITPβ have specific roles in cells. In recent years, new functions
for PITPs have been described which include roles in neurite outgrowth [31],
cytokinesis [32] and in regeneration and function of stem cells [33].
PITPα is enriched in the brain [25]. This observation, coupled with the
phenotype of PITPα knockout mice, which suffer from neurodegenerative
disorders, suggest that PITPα contributes to the functioning of the nervous
systems in mammals. In a recent study, it was demonstrated that PITPα plays
a role in neurite extension. Neurite extension is essential for wiring of the
nervous system during development, and one extracellular guidance cue that
promotes axonal growth is a family of secreted proteins collectively called
netrins. The netrin receptors, DCC (deleted in colorectal cancer) and neogenin
in vertebrates, are conserved across species (e.g. UNC40 in C. elegans and
Frazzled in Drosophila). In vertebrates, netrin-1-induced neurite outgrowth
involves an interplay of many signalling cascades including the Rho family of
GTPases, PLCγ, phosphoinositide 3-kinase, ERK (extracellular-signal-regulated
kinase) and FAK (focal adhesion kinase ) [34,35]. Xie et al. reported that PITPα
(but not PITPβ) interacts with the netrin receptors, DCC and neogenin, and
that this interaction is mediated by the C-terminus of PITPα, which is distinct
from the C-terminus of PITPβ. Netrin-1 was shown to increase tyrosine
phosphorylation of PLCγ and also increased its catalytic activity, resulting in
PtdIns(4,5)P2 hydrolysis. Netrin-induced neurite outgrowth was impaired in
mice with reduced PITPα expression, as was netrin-1-stimulated PLC activity.
In addition, netrin-1-stimulated neurite outgrowth was inhibited by an inhibitor
of PLC. Collectively, these data suggest that PLCγ and PITPα are required for
neurite outgrowth when neurons are stimulated with netrin-1.
The cell division cycle culminates in cytokinesis, the process that follows
duplication and spatial segregation of chromosomes [36]. In animal cells, an
actomyosin contractile ring constricts the plasma membrane, forming a membrane furrow; this is accomplished by delivery of new membranes [37–41].
Ingression terminates when the furrow reaches the spindle mid-zone [36,38].
For the final separation of cells, the contractile ring and the spindle mid-zone
disassemble and the intercellular bridge between the daughter cells is severed
[37]. In Drosophila, the single Class I PITP, Gio, is required for both mitotic and meiotic cytokinesis. Gio is involved in actomyosin ring constriction.
In cells mutated for Gio, an abnormal accumulation of Golgi-derived vesicles
was observed, suggesting the failure of these vesicles to fuse with the invaginating furrow membrane. Local production of PtdIns(4,5)P2 plays a crucial role in
completion of cytokinesis [42–44] and its depletion blocks cytokinesis [43,45].
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In Drosophila and Schizosaccharomyces Pombe, mutations in a PtdIns4Kβ also
cause a defect in cytokinesis [46,47]. Analysis of PtdIns4Kβ mutants also revealed an abnormal accumulation of Golgi vesicles, resulting in defects in constriction of the actomyosin ring, thereby suggesting that PITP and PtdIns4Kβ
might function in the same pathway. The function of PtdIns(4,5)P2 in cytokinesis is probably complex because it can participate in vesicle delivery and
regulation of actin dynamics, as well as being the substrate for PLC. One
study suggested that PtdIns(4,5)P2 mediates contact between the contractile ring and the plasma membrane [43], whereas two other studies suggested
that PtdIns(4,5)P2 hydrolysis by PLC was required for cytokinesis [48,49].
In mammalian cells, PITPβ is potentially essential for cytokinesis based on
its localization. Whereas in interphase cells, PITPβ localizes to the Golgi, in
metaphase it is found along microtubules between the opposing spindle poles.
At anaphase and telophase, PITPβ is concentrated on the central spindle, at
late telophase it localizes at the mid-zone and during late stage of cytokinesis
PITPβ is detected on the interconnecting cytoplasmic bridge (N. Carvou and
S. Cockcroft, unpublished work).
The function of PITP in cytokinesis is analogous to that of the unrelated
yeast PITP, Sec14p, in S. pombe. Sec14p is required for both membrane delivery
to assemble the forespore membrane [50] and structural integrity of the spindle
pole body during meiosis [51]. Another Sec14 protein, patellin-1, is also required
during cytokinesis in plants. Patellin-1 localizes to the cell plate and is involved in
membrane trafficking events associated with cell-plate expansion [52]; its ability to
bind to phosphoinositides appears to be essential for patellin-1 function. Finally,
RdgBαI (Nir2), which contains a PITP domain (Table 1), is phosphorylated
during mitosis and is also required for completion of cytokinesis [53].
Planarians are bilaterally symmetric metazoans renowned for their
regenerative capacities associated with their neoblasts, a pluripotent adult stem
cell population [54]. Neoblasts are the only mitotically active cells in planarians
and can give rise to 40 different cell types found in the adult organism. In intact
planarians, stem cells (neoblasts) replace cells lost to normal physiological turnover,
and in amputated animals, to the regeneration blastema, the structure in which
missing tissues are regenerated. Using an RNAi (RNA interference) screen, 240
genes were identified that are required for regeneration, tissue homoeostasis and
stem cell regulation. The gene encoding PITP was found to be required for basal
neoblast functioning in regeneration. Additionally, RNAi knockdown of PITP
also has a decreased number of mitotic cells, suggesting defects in cell division.
This requirement for PITP in mitosis is reminiscent of a PITP requirement seen
during cytokinesis in Drosophila (described above), underlining a very basic
function fulfilled by PITP in cell division. In mammalian cells, where there are two
Class I PITPs, our studies suggest that it is PITPβ that is essential for cell division.
Concluding remarks
PITP is beautifully designed to encapsulate a lipid and its most favoured
lipid cargo is PtdIns. Thus proteins of the PITP family are most likely to fulfill
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functions that lie at the interface of phosphoinositide-based lipid signalling
and/or lipid metabolism. The possibility that some PITPs participate in
PtdCho-based signalling cannot be excluded, however [55]. The underlying
theme appears to be the ability of PITPs to manipulate local lipid levels to
form an appropriate lipid environment whether for signalling purposes or
for membrane delivery. Both signal transduction and membrane delivery are
primary events required for many aspects of biology, including cell division,
transport of secretory products and expansion of membranes for cell growth
(e.g. neurite outgrowth). PITPs are implicated in many of these events.
Despite our increased knowledge of their biological requirements, a detailed
understanding of how the lipid transfer activity is harnessed by the cell is still
lacking. The availability of structural information will allow specific questions,
such as what is the significance of PtdIns and PtdCho binding and delivery,
to be addressed. PITP mutants specifically deficient in PtdIns binding and
transfer can now be made for PITPs from different species and should provide
the answer to the ultimate question of whether it is the essential role of PITPs
in living cells to bind and transport PtdIns.
The work in the Cockcroft Laboratory is supported by the Wellcome Trust. I thank
Judith Murray-Rust for preparing Figure 1B and Sadaf Shadan for her comments on
the manuscript.
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