POLYPRENOL REDUCTASE2 Deficiency Is Lethal in Arabidopsis Due to Male Sterility

Abstract

Dolichol is a required cofactor for protein glycosylation, the most common posttranslational modification modulating the stability and biological activity of proteins in all eukaryotic cells. We have identified and characterized two genes, PPRD1 and -2, which are orthologous to human SRD5A3 (steroid 5a reductase type 3) and encode polyprenol reductases responsible for conversion of polyprenol to dolichol in Arabidopsis thaliana. PPRD1 and -2 play dedicated roles in plant metabolism. PPRD2 is essential for plant viability; its deficiency results in aberrant development of the male gametophyte and sporophyte. Impaired protein glycosylation seems to be the major factor underlying these defects although disturbances in other cellular dolicholdependent processes could also contribute. Shortage of dolichol in PPRD2-deficient cells is partially rescued by PPRD1 overexpression or by supplementation with dolichol. The latter has been discussed as a method to compensate for deficiency in protein glycosylation. Supplementation of the human diet with dolichol-enriched plant tissues could allow new therapeutic interventions in glycosylation disorders. This identification of PPRD1 and -2 elucidates the factors mediating the key step of the dolichol cycle in plant cells which makes manipulation of dolichol content in plant tissues feasible.

Modeling of Dolichol Mass Spectra Isotopic Envelopes as a Tool to Monitor Isoprenoid Biosynthesis

“…One is also tempted to speculate that methylerythritol cyclodiphosphate, a product of the MEP pathway so far recognized as a retrograde signaling metabolite inducing the expression of selected abiotic stressresponsive genes (Xiao et al, 2012), could be involved. Notably, methylerythritol cyclodiphosphate potentiates the unfolded protein response (Walley et al, 2015) induced upon osmotic stress in plants (Moreno and Orellana, 2011), and interestingly, Dol attenuates this stress reaction (Jozwiak et al, 2015).…”

“…Accordingly, a mild increase in the level of PPRD2 mRNA (Fig. 1F), the main PPRD isoform responsible for the conversion of Prens to Dol in Arabidopsis seedlings (Jozwiak et al, 2015), was observed.…”

“…CPT or PPRD2 expression analysis was performed in a total volume of 20 mL of Luminaris Color HiGreen High ROX qPCR Master Mix (Thermo Scientific) using gene-specific primers (Jozwiak et al, 2013(Jozwiak et al, , 2015 in the StepOnePlus Real-Time PCR System (Applied Biosystems) according to the manufacturer's instructions.…”

“…Their biosynthesis is catalyzed by cisprenyltransferases (CPTs), and six out of nine putative CPT-encoding genes of Arabidopsis are expressed in roots (Surmacz and Swiezewska, 2011). Eukaryotic CPT performs sequential condensations of the IPP with an all-trans-initiator to form a mixture of homologous polyprenyl diphosphates, which subsequently are converted to Dols according to tissue-specific requirements (Cantagrel et al, 2010;Jozwiak et al, 2015). Structural analysis of polyisoprenoids of various species CPTs from S. cerevisiae are Rer2 and Srt1 (P35196 and Q03175); those from Arabidopsis are AtCPT1, AtCPT2, AtCPT3, AtCPT6, AtCPT7, and AtCPT9 (O80458, F4IMI8, Q8S2T1, Q8RX73, Q56Y11, and Q570Q8).…”

Polyprenols Are Synthesized by a Plastidial cis-Prenyltransferase and Influence Photosynthetic Performance

“…This search yielded two candidates: AtCPT2 (At2g23400) and AtCPT7 (At5g58770). While both CPTs are expressed in leaves, AtCPT2 is absent in stem tissue, which is known to contain polyprenols (Surmacz and Swiezewska, 2011;Jozwiak et al, 2015); hence, AtCPT7 emerged as the most promising candidate. To explore the involvement of AtCPT7 in polyprenol synthesis, we first isolated a single homozygous T-DNA line for the gene locus At5g58770 (SALK_022111; Figure 1A) and then generated three independent RNAi lines targeting AtCPT7 for mRNA knockdown as well as transgenic lines overexpressing AtCPT7 (CPT7-OE).…”