# Order in Chaos: Lesser-Conserved and Repeat Structures in Dehydrins
## 3 Results
### Dhn Groups and Orders  
The BLASTp searches on the 2 variants of the K segment yielded 8675 unique candidate sequences (candidate sequences csv file, Supplementary File S1). After the initial assessment of the results with lower threshold values, the threshold scores for the recognition of K, H, F, Y, and S segments were set at 6.0, 6.8, 5.0, 3.5, and 5.2, respectively. Using these values, the Dhn Decoder R script identified K-like segments in 4044 sequences, including 293 designated as partial according to the NCBI description or by lack of an N-terminal M residue. A total of 1142 sequences were reclassified as pseudo Dhns or reticulons (discussed below), leaving 2658 complete and 236 partial sequences with well-conserved K segments broadly distributed across green plant groups (Table 1). The resulting list still includes some likely paralogues or allelic variants in some taxa, often differing by indels of sufficient length to result in <90% identity, and it may exclude some known Dhns with >90% identity with Dhns in the final list. The K1, Kn, KnS, and irregular groups included 982 sequences with 1 or 2 single, relatively weakly conserved K segments, with consensus residues in some positions but others not conforming to type. Because these weak segments leave some doubt as to whether these are functional Dhns, all Dhns in these four groups with maximum K segment scores <7.0 were given a provisional status as pseudo Dhns. After this screening, the remaining K1 Dhns were folded into the Kn group in further analysis. The search algorithm also identified a group of 160 KS sequences with a fused KS-like segment (e.g., KEESLMEKIKEKLLHGDSSSSD, NCBI accession XP_039115618.1, _Dioscorea cayensis_) near the N-terminus rather than with a C-terminal S segment as in typical (H)KnS Dhns, and with the K segment portion usually scoring < 7.0. Some of these were tagged as reticulons in the NCBI database. Known Arabidopsis reticulons contain a similar KS-like segment near the N-terminus [20]. Most of these were already identified in NCBI as reticulons or reticulon-like; additional putative reticulons were identified manually by screening all KS DHNs identified by the script for lack of a terminal S segment and then checking for the presence of the fused KS-like segment near the N terminus. These sequences are well-conserved across a broad range of land plants (Figure S1). Sequences reclassified as reticulons and pseudo DHNs were excluded from further analysis but included in the database because they may be of future interest. This raises the possibility that anti-K segment antibodies may bind to reticulons and the low-scoring K segments in pseudo Dhns, resulting in misidentification of these proteins as Dhns using immunological methods. Although eukaryote reticulons are recognized by a conserved reticulon homology \begin{table}
\begin{tabular}{l c c c c c c} \hline \hline  & \multicolumn{6}{c}{**Dhn Group or Order**} \\ \hline
**Taxon** & **Kn** & **SKn** & **HKn(S)** & **F(S)Kn** & **Yn(S)Kn** & **Irr** & **Total** \\ \hline Algae & 5 & 2 & & & & 7 \\ Bryophyte & 18 & & & 5 & 4 & 27 \\ Lycophbyte & & 3 & 2 & & & 5 \\ Fern & 3 & & 6 & & & 9 \\ Gymnosperm & 26 & 8 & & 36 & & 3 & 73 \\ Basal angiosperm & 1 & 3 & 1 & 1 & 2 & 8 \\ Magnoliid & & 2 & 2 & 6 & 7 & & 17 \\ Monocot & 92 & 91 & 54 & 45 & 232 & 16 & 530 \\ Basal eudicot & 13 & 6 & 17 & 24 & 26 & 1 & 87 \\ Caryophyll & 3 & & & 22 & 16 & 1 & 42 \\ Rosid & 127 & 40 & 266 & 302 & 448 & 23 & 1206 \\ Asterid & 76 & 75 & 107 & 154 & 228 & 7 & 647 \\ \hline Grand total & 364 & 230 & 455 & 590 & 964 & 55 & 2658 \\ \hline \hline \end{tabular}
\end{table} 
Table 1: Distribution of Dhn types among phylogenetic groups for 2658 complete Dhn sequences identified by Dhn Decoder. domain at the C-terminus (Figure S1), it is noteworthy and perhaps intriguing that plant reticulons have this Dhn-like characteristic. Dhn Decoder identified 69 GC-variant F segments among the 620 F(S)Kn Dhns, with 17 in asterid species and the remainder in rosids.