Auxin as an intercellular signal - Intercellular Communication in Plants - page 8

and found to encode a 622 amino acid protein with up to 12 putative transmembrane
segments with similarity to a group of transporters from bacteria (Galweiler et al .,
1998). Simultaneously, a homologous gene was identified independently by several
groups - the PIN2 / EIR1 / AGR1 gene (Chen et al ., 1998; Luschnig et al ., 1998; Muller
et al ., 1998; Utsuno et al ., 1998) - and analysis of additional homologs ( PIN3 , PIN4
and PIN7 ) followed (Friml et al. , 2002a,b, 2003). In total, the Arabidopis PIN gene
family consists of eight members and homologous genes were found in other plant
species, e.g. maize, rice, soybean and others. The proposed function for PIN pro-
teins as efflux carriers has not ultimately been proven; nonetheless, several lines of
evidence strongly support their role in PAT:
1. Topology and localization of PIN proteins: The PIN proteins share more
than 70% similarity and have almost identical topology-alarge hy-
drophilic loop is symmetrically flanked by two conserved, highly hy-
drophobic domains with five to six transmembrane segments. Transporters
of the major membrane facilitator class display similar topology (Chen
et al ., 1998; Luschnig et al ., 1998; Muller et al ., 1998; Utsuno et al .,
1998). When localized in planta , most PIN proteins show asymmetric cel-
lular localization (Plate 1.1B-D), impressively correlating with the known
direction of PAT in these tissues. This polar localization was predicted by
chemiosmotic hypothesis for auxin efflux proteins (Rubery & Sheldrake,
1974; Raven, 1975).
2. Heterologous expression of PIN proteins: To date, the only experimental
system used to address PIN transport activity is yeast assay (Luschnig et al. ,
1998). Yeast carrying a mutation in the GEF1 gene (resulting in an altered
ion homeostasis) shows enhanced resistance to the yeast toxin fluoroindole,
when overexpressing PIN2/EIR1/AGR1. Fluoroindole shows some (albeit
limited) structural similarity to auxin (Luschnig et al ., 1998). The yeast
also retains less radioactively labeled auxin than does control yeast (Chen
et al. , 1998). Nonetheless, measurements of auxin efflux instead of auxin
retention have not been demonstrated so far.
3. PAT is defected in pin mutants: All defects observed so far in pin mutants
occur in processes known to be regulated by PAT and they can be pheno-
copied by treatment of wild-type plants with AEIs. One of the strongest
arguments for the involvement of PIN proteins in auxin transport is a
reduction of PAT in pin mutants, which directly correlates with loss of
PIN expression in corresponding tissue, as was demonstrated for basipetal
auxin transport in stem of pin1 mutant or in root of pin2 mutant (Okada
et al ., 1991; Rashotte et al ., 2000). In addition, local distribution and ac-
cumulation of auxin monitored both by the activity of an auxin responsive
construct (e.g. DR5::GUS ; Sabatini et al ., 1999) and by direct measure-
ments of auxin content (Friml et al ., 2002b) or using radioactive IAA
preloaded root tips (Chen et al ., 1998) have been shown to be affected in pin
mutants.