Salt tolerance of Arabidopsis thaliana requires maturation of N -glycosylated proteins in the Golgi apparatus

Abstract

Protein N -glycosylation in the endoplasmic reticulum (ER) and in the Golgi apparatus is an essential process in eukaryotic cells. Although the N -glycosylation pathway in the ER has been shown to regulate protein quality control, salt tolerance, and cellulose biosynthesis in plants, no biological roles have been linked functionally to N -glycan modifications that occur in the Golgi apparatus. Herein, we provide evidence that mutants defective in N -glycan maturation, such as complex glycan 1 ( cgl1 ), are more salt-sensitive than wild type. Salt stress caused growth inhibition, aberrant root-tip morphology, and callose accumulation in cgl1 , which were also observed in an ER oligosaccharyltransferase mutant, staurosporin and temperature sensitive 3a ( stt3a ). Unlike stt3a , cgl1 did not cause constitutive activation of the unfolded protein response. Instead, aberrant modification of the plasma membrane glycoprotein KORRIGAN 1/RADIALLY SWOLLEN 2 (KOR1/RSW2) that is necessary for cellulose biosynthesis occurred in cgl1 and stt3a . Genetic analyses identified specific interactions among rsw2 , stt3a , and cgl1 mutations, indicating that the function of KOR1/RSW2 protein depends on complex N -glycans. Furthermore, cellulose deficient rsw1-1 and rsw2-1 plants were also salt-sensitive. These results establish that plant protein N -glycosylation functions beyond protein folding in the ER and is necessary for sufficient cell-wall formation under salt stress.

Role of complexN-glycans in plant stress tolerance

“…Importance of functional cellulose biosynthesis for salt tolerance was further supported by the novel finding of increased salt-sensitivity of rsw2-1 and rsw1-1 single mutants. 1 Our previous and current data have implications that affect our view of protein N-glycosylation in plants. First, after all, plant complex N-glycans confer important in vivo functions to secreted/ secretory glycoproteins, i.e., protect root growth from salt/osmotic stress.…”

“…This showed that cgl1 and stt3a enhance cellulose deficiency of rsw2-1, and in turn indicate that the KOR1/RSW2 protein requires complex N-glycans for its function in vivo. Further pyramiding of these mutations resulted in incremental enhancement of growth defects as well as developmental defects of the host plants (Kang et al, (2008), and Fig. 1).…”

“…We recently reported that mutants defective in complex N-glycans show enhanced salt sensitivity, establishing that complex N-glycans are indispensable for certain biological functions. 1 Our previous study using an OST subunit mutant stt3a indicated that protein glycosylation could affect salt tolerance and root growth of A. thaliana. 14 Since OST functions upstream of protein folding processes in the ER, stt3a caused an unfolded protein response (UPR), which is a general ER stress response to protein folding defects, as well as accumulation of under-glycosylated proteins.…”

“…1 A temperature sensitive rsw2-1 allele 16 showed specific genetic interaction with both cgl1 and stt3a mutations. The corresponding double mutants exhibited spontaneous growth defects at the permissive temperature that were reminiscent of those of rsw2-1 at the restrictive temperature, of cgl1 and stt3a plants treated with salt, and of the rsw1-1 rsw2-1 double mutant that combines two cellulose deficiency mutations.…”

“…In our recent study, we tried to address whether the salt stress response of the mutant is caused by an activation of UPR, or by a shortage of functional glycoproteins produced by the cells. 1 The cgl1 mutant is defective in N-acetylglucosaminyltransferase in the Golgi apparatus 15 and only able to produce oligomannosidic-type N-glycans but not complex-type N-glycans. 8 cgl1 mutants exposed to salt stress exhibited root growth arrest and radial swelling similar to stt3a mutants, however, unlike stt3a, the cgl1 mutation did not cause UPR as judged by expression of an UPR marker gene, BiP pro -GUS.…”