Plant Molecular Biology 36: 541–552, 1998.
1998 Kluwer Academic Publishers. Printed in Belgium.
Changes in the levels of seven proteins involved in polypeptide folding and
transport during endosperm development of two barley genotypes differing
in storage protein localisation
& David J. Simpson
Department of Physiology, Carlsberg Laboratory. Gamle Carlsbergvej 10, 2500 Valby, Denmark (
Present address: Department of Biochemistry, University of Cambridge, Tennis Court Road,
Cambridge CB2 1QW, UK
Received 7 August 1997; accepted in revised form 8 October 1997
Key words: endoplasmic reticulum, chaperones, storage protein accumulation, transport mutant, Hordeum vulgare
The Russian barley cultivar Nevsky lacks
hordein and accumulates most of its hordein in the lumen of the
endoplasmic reticulum and only a minor portion in the vacuole. In wild type barley and all other temperate cereals,
storage proteins are deposited in the vacuole. F1 crosses revealed that the Nevsky phenotype is recessive; but the
extent of hordein accumulation in the endoplasmic reticulum in F2 endosperm lacking
hordein was very much
less than in the Nevsky parent. In order to study the Nevsky endosperm phenotype we have measured the levels
of seven proteins and two mRNAs involved in protein folding in the ER lumen or ER to Golgi transport during
endosperm development. The protein levels were unaltered in Nevsky as compared to the wild-type variety Bomi.
Whenthelevels oftheseseven proteinswerecorrelatedwith the rate of hordeinaccumulation, four ofthese (HSP70,
PDI, Sar1p and Sec18p) were consistently up-regulated with hordein synthesis. Accumulation of hordein in the
endoplasmic reticulum appears to be determined by the absence of
hordein, or the product of a gene closely
linked to it, plus one or more other recessive genes.
Prolamins, the major protein reserves of cereal grains,
are synthesised and deposited in the developing endo-
sperm. In temperate cereals, prolamins are transpor-
ted to the vacuole, whereas in tropical cereals they
accumulate in the lumen of the endoplasmic reticulum
(ER). In barley, hordeins are exclusively deposited in
the vacuole [5, 6], but the fully viable Russian bar-
ley cultivar Nevsky, which is unusual in lacking
hordein, accumulates most of its hordeins in lumen
of the ER, and thus resembles tropical cereals .
However, in the subaleurone layer of Nevsky, where
endosperm,all hordeinsare transportedto the vacuole.
This indicates that the rate of protein synthesis may
determine the extent of hordein deposition in the ER,
rather than its unusual hordein composition .
Hordeins are synthesised by ER-bound ribosomes
on the surface of the rough ER and are cotranslation-
ally transported into the lumen of the ER, where a set
of resident proteins assists folding of the nascent poly-
peptides . Once the hordeins have obtained their
native conformation they are exported to the vacuole
via the Golgi apparatus (Golgi) and the prevacuolar
compartment by vesicle-mediated transport [10, 30,
33, 47]. Due to difﬁculties in monitoring transport of
these non-glycosylated prolamins through the Golgi,
there has been some controversy about whether pro-
lamins go via the Golgi, directly from the ER to the
vacuole, or by both pathways . In barley there is
no evidence of a direct ER to vacuole pathway in the
developing endosperm  and recently hordeins have
beenshown in theGolgi apparatusbyimmunoelectron
microscopy (D.J. Simpson, unpublished).
The sulfur-rich B and
hordeins, which account
for ca. 80% of the total hordein fraction, can form