Characterization and expression of the Douglas-fir luminal binding protein
Bibliographic record
Abstract
The endoplasmic reticulum (ER) molecular chaperone, BiP, plays a role in the translocation and subsequent folding and assembly of newly synthesized proteins targeted to the ER and secretory pathway. The sequence encoding a Douglas-fir (Pseudotsuga menziesii [Mirb] Franco) BiP homologue (PmBiP) was identified by differential screening of a seedling cDNA library. Southern blotting indicated that PmBiP is most likely present as a single copy although other BiP alleles likely exist within a given seedlot. The deduced amino acid sequence of PmBiP contains a HEEL tetrapeptide sequence which functions to retain PmBiP in the ER and is different from HDEL commonly found in angiosperm plant BiPs. Amino acid sequence alignment and phylogenetic analysis show that PmBiP is highly similar to other plant BiPs yet forms a distinct phylogenetic subgroup separate from angiosperm BiPs. Northern and western blotting revealed that PmBiP is subject to developmental regulation during seed development, germination, and early seedling growth and is seasonally regulated in needles of young seedlings. The expression of PmBiP is developmentally regulated during seed development with higher amounts present in seeds prior to embryo development and the deposition of storage proteins. Increased PmBiP expression correlates with seedling growth and developm ent and the mobilization of seed storage proteins. Increased synthesis during germination is likely due to increased synthesis of cell wall proteins and other secretory traffic. This idea is supported by immunolocalization of PmBiP in root tip cells showing staining around the new cell wall in telophase cells and at the periphery of cells in the elongation zone. PmBiP may also play a role in mediating homotypic ER and nuclear envelope membrane fusion during mitosis in actively dividing tissues. PmBiP is seasonally regulated in the needles of young seedlings and increased expression was observed in tissues treated with low temperature suggesting that PmBiP plays an important role in the adaptation of seedlings to low temperatures. This is most likely accomplished through the maintenance of secretory traffic through the ER necessary for the synthesis of proteins with a more direct role in cold acclimation. Proteins were associated with PmBiP in an ATP dependent manner in mature seeds and 2-day-old seedlings but were only detectable in minute amounts. ATP associated proteins were more readily detectable in embryonal suspensor mass (ESM) cultures but only in small amounts unsuitable for N-terminal sequencing and identification. The Douglas-fir BiP promoter (PmBiPProl) contains a variety of cis-acting regulatory elements commonly found in the promoters of storage protein genes, light regulated genes, and phenlypropanoid and cell wall protein genes. The presence of different cis-element groups suggests the transcriptional regulation of PmBiP is controlled by a variety of signal transduction pathways depending upon the developmental and/or physiological state of a given tissue. Transient expression analysis showed that PmBiPProl is functional in germinating Douglas-fir embryos. The expression of PmBiPProl in transgenic Arabidopsis is associated with actively dividing and secretory tissues. Deletion analysis showed that minimal promoter elements lie within a 263 bp region directly upstream of the PmBiP cDNA sequence although upstream flanking sequences are necessary for higher level expression. G-box motifs residing within the 263 bp fragment together with a quantitative activator region (QAR) and a negative regulatory region (NRR) present in upstream areas are likely involved in transcriptional control in young seedlings. PmBiPProl was also wound inducible in transgenic Arabidopsis cotyledons that correlated with similar experiments conducted in Douglas-fir seedlings. Elements involved in conferring wound inducibility are located in PmBiPProl-5 but upstream elements are necessary for higher level expression. G-box motifs may also play a role in the wound inducibility of the Douglas-fir BiP promoter.
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How this classification was reachedexpand
Full frame machine prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. The Gemma side is a direct model label for every work in the frame, read from the title-only record. The Codex side is a classifier learned from the 10,348 direct Codex labels and calibrated to design-weighted sample rates; fields without enough sample support carry no Codex call. Candidate is the union of the two sides; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels.
Distilled classifier scores by category (both heads)
| Category | Codex | Gemma |
|---|---|---|
| Metaresearch | 0.000 | 0.000 |
| Meta-epidemiology (narrow) | 0.000 | 0.000 |
| Meta-epidemiology (broad) | 0.000 | 0.000 |
| Bibliometrics | 0.000 | 0.000 |
| Science and technology studies | 0.000 | 0.000 |
| Scholarly communication | 0.000 | 0.000 |
| Open science | 0.000 | 0.000 |
| Research integrity | 0.000 | 0.000 |
| Insufficient payload (model declined to judge) | 0.000 | 0.001 |
Machine scores (provisional)
The two teacher heads of the student model, read on this work. A score orders the frame for review; it never asserts a category, and the validation status ships verbatim with every row.
Baseline scores from an immature model (maturity gate not passed, 7 training rounds). Scores rank; they never assert a category.
score_only:v0-immature-baseline · verbatim from the scoring run: score_only means the number may rank works, and no category label ships from itClassification
machine, unvalidatedMachine predicted; a candidate call from one source (direct Gemma or distilled Codex), not a consensus.
How this classification was reached, model by model and score by score, is at the end of the page under "How this classification was reached".