Plant Science 157 2000 1 – 12
Lipid transfer proteins are encoded by a small multigene family in Arabidopsis thaliana
Vincent Arondel
1 2
, Chantal Vergnolle
2
, Catherine Cantrel, Jean-Claude Kader
Laboratoire de Physiologie Cellulaire et Mole´culaire, CNRSUni6ersite´ Pierre et Marie Curie UMR
7632
, Case
154
,
4
Place Jussieu, F-
75252
Paris cedex
05
, France Received 26 October 1999; received in revised form 28 January 2000; accepted 21 February 2000
Abstract
Lipid transfer proteins LTPs are small, basic and abundant proteins in higher plants. They are capable of binding fatty acids and of transferring phospholipids between membranes in vitro. LTPs from this family contain a signal peptide and are secreted
in the cell wall. Their biological function is presently unknown. LTPs have been suggested to participate to cutin assembly and to the defense of the plants against pathogens. A genetic approach should prove useful to provide clues on their in vivo functions.
Here, the characterization of the LTP gene family in Arabidopsis thaliana is described. At least 15 genes were identified, their map position determined and the expression pattern characterized for six of them. All the sequences exhibit the typical features of plant
LTPs. The molecular weight is close to 9 kDa, the isoelectric point is near 9 except for three acidic LTPs, and typical amino acid residues such as cysteines are conserved. Genomic DNA blotting hybridization experiments performed using ltp1 to ltp6 as
probes indicate that ltps form distinct 1 – 3 gene subfamilies which do not cross hybridize. Expression studies indicate that all the genes tested are expressed in flowers and siliques, but not in roots. Ltp1, ltp5 and ltp2 are expressed significantly in leaves, while
ltp6 is detected only in 2 – 4-week-old leaves. In addition, ltp4 and ltp3 are strongly upregulated by abscisic acid ABA. Tandem repeats can be noted concerning ltp1 and ltp2 on chromosome 2, ltp3 and ltp4 on chromosome 5 and ltp5 and ltp12 on
chromosome 3. While ltp7, ltp8 and ltp9 map at the same position on chromosome 2, the other genes are dispersed throughout the genome. The characterization of the Arabidopsis ltp gene family will permit to initiate a genetic approach for determining the
in vivo functions of these proteins. © 2000 Elsevier Science Ireland Ltd. All rights reserved.
Keywords
:
Lipid transfer protein; Arabidopsis thaliana; cDNA; Tandemly repeated genes; Abscisic acid www.elsevier.comlocateplantsci
1. Introduction
Proteins capable of transferring lipids between membranes in vitro have been purified from a
wide range of living organisms [1]. Some of them have been cloned and the amino acid sequence
comparisons revealed that these proteins fall into several different classes that are unrelated based
on their primary structure. Although all these proteins, called lipid transfer proteins LTP, were
initially supposed to participate to membrane bio- genesis, no clear evidence of such a role has been
demonstrated in vivo. Actually, the biological function of some LTPs [2] begins to be investi-
gated. For example, the sec14 protein of yeast, which is a phosphatidylinositol transfer protein
Abbre6iations
:
ABA, abscisic acid; BAC, bacterial artificial chro- mosome; BLAST, basic local alignment search tool; EST, expressed
sequence tag; LTP, lipid transfer protein; NMR, nucleic magnetic resonance; PCR, polymerase chain reaction; PITP, phosphatidylinosi-
tol transfer protein; RFLP, restriction fragment length polymor- phism; TAIR, The Arabidopsis Information Resource; T-DNA,
transferred DNA; TIGR, The Institute for Genomic Research; YAC, yeast artificial chromosome.
3 Accession numbers: ltp1: AF159798; ltp2: AF159799; ltp3: AF159800; ltp4: AF159801; ltp5: AF159802; ltp6: AF159803.
Corresponding author. E-mail address
:
kaderccr.jussieu.fr J.-C. Kader.
1
Present address: Laboratoire de Lipolyse Enzymatique, UPR CNRS 9025, Universite´ de la Me´diterrane´e, Marseille, France.
2
These authors have contributed equally to this work 0168-945200 - see front matter © 2000 Elsevier Science Ireland Ltd. All rights reserved.
PII: S 0 1 6 8 - 9 4 5 2 0 0 0 0 2 3 2 - 6
PITP was shown to be a sensor of the lipid composition of the Golgi membrane, and its ca-
pacity to
down regulate
phosphatidylcholine biosynthesis in this organelle was demonstrated. In
addition, mammalian PITPs, which are struc- turally unrelated to sec14p, seem to play a key role
in phospholipase C mediated signaling through their
binding capacity
to phosphoinositides.
Therefore, it appears that these two different LTPs do not possess identical physiological functions
and that neither of them transfer lipids between intracellular membranes.
In higher plants, LTPs form a very homoge- neous class of protein, if a sec14-like PITP is
excluded [3]. They are small 9 kDa, abundant and basic proteins that contain eight cysteine
residues [4,5]. They are capable of transferring several different phospholipids, and they can bind
fatty acids [6] and acyl-CoA esters. Structural data have been recently published, based on both X-ray
diffraction [7] and nucleic magnetic resonance NMR [8] techniques. These results indicate that
LTPs contain a hydrophobic pocket capable to accommodate a fatty acid or a lysophospholipid
molecule.
Numerous LTP cDNAs have been cloned from different plant species [4]. These data have indi-
cated the existence of multiple isoforms, that are differently expressed and regulated [9 – 17]. How-
ever, most of these genes are preferentially ex- pressed in epidermal cells of leaves and in flowers,
and very rarely in roots.
All non-specific plant LTPs characterized so far contain a signal peptide, and immunolocalization
data indicate that they locate to the cell wall [18]. These proteins have also been shown to be
secreted by cell cultures [15,19]. This localization therefore preclude a priori an intracellular role for
these proteins. Possible biological functions have been suggested. LTP might play a role in cutin
and wax assembly [20,21]. Another possible role is based on the antifungal properties displayed by
some LTP [22]. These proteins might play a role in the defense of the plant against pathogen attack
[23 – 25]. Indeed, it has been shown that increasing the level of an LTP in transgenic tobacco enhances
the resistance of the plant towards a pathogen [26]. A possible way to find a role for these
proteins would consists in obtaining mutants or transgenic plants that express antisense RNA. The
phenotypic characterization of these plants would provide clues with regards to the in vivo function
of these proteins. Arabidopsis thaliana seems to be the most appropriate plant material for a genetic
approach, since it is very easy to transform [27], that numerous tools are available that allows re-
verse genetics transferred DNA, T-DNA, [28] or transposons tagged lines and that the genome
programs have yielded a considerable amount of genomic and cDNA sequences [29,30]. Here, the
characterization of the Arabidopsis ltp gene family is described.
2. Material and methods
