Volume 8 issue 10 October 2007
Localization of Vacuolar Transport Receptors and Cargo Proteins in the Golgi Apparatus of Developing Arabidopsis Embryos
Giselbert Hinz, Sarah Colanesi, Stefan Hillmer, John C. Rogers, and David G. Robinson

Supplemental Figure 1: Expression of cruciferin and aleurain during Arabidopsis development. Expression of cruciferin ( At4g28520) and aleurain (At5g60360) were analyzed during plant development employing the AtGenExpress Visualization tool. The result is plotted as normalized mean intensity.

Figure 1 (.jpg)

Supplemental Figure 2: Alignment of the family of aleurain-homologous cysteine proteases. The sequence alignment was performed using the Clustawl program (57) provided by the EMBL-EBI database and search engine. Accession numbers for the sequences were as follows:
Hordeum vulgare Aleurain BAF02548; Hordeum vulgare EP-B P25250; Lolium multiflorum Cystein proteinase CAB71032; Oryza sativa oryzain gamma chain precursor P25778; Zea mays cystein protease CAA68192; Zea mays cystein protease 2 Q10717; Brassica napus cystein protease ABA71255; Nicotiana tabacum cystein protease BAA96501; Brassica oleracea senescence-associated cystein protease AAL60582; Nicotiana benthamiana cystein protease aleurain type AAZ32410; Nicotiana tabacum NTCP23-loke cystein protease AAK07729; Pisum sativum cystein protease ABF18679; Pisum sativum early leaf senescence abundant cystein protease CAC41636; Solanum lycopersicum cystein proteinse 3 precursor Q40143; Arabidopsis thaliana thiol protease aleurain precurser AtAleu Q8H166, At5g60360; Arabidopsis thaliana thiol protease aleurain-like precursor Q8RWQ9, At3g45310.
A) Alignment of aleurain family proteins.
B) Alignement of Hordeum vulgare aleurain BAF02548 (Aleurain) and EP-B P25250 (EP-B)
The conserved NPIR/NPIK ssVSD is printed in red. The conserved C-terminal hydrophobic putative ctVSD is printed in green. The 127 C-terminal sequence of Arabidopsis aleurain At5g60360 which was used as antigen for immununization is shaded in grey. Asterisks indicate identical amino acids, double points are conserved exchanges, single points are homologoue amino acids.

Figure 2 a (.jpg)

Figure 2 b (.jpg)

Supplemental Figure 3: Sequences of aspartic protease A1 and beta VPE. The sequences of the aspartic protease A At2g11910 and beta vacuolar processing enzyme At1g62710. Putative N-terminal ssVCD is printed in red, putative hydrophobic C-terminal ctVSD are printed in green.

Figure 3 (.jpg)

Supplemental Figure 4: Alignment of putative antigenic cysteine proteases in Arabidopsis seeds.
(A) C-terminal 127 amino acids of Arabidopsis thaliana cysteine protease aleurain precursor AtAleu Q8H166, At5g60360.
(B) Cysteine proteases bearing a putative hydrophobic C-terminal ctVSD were aligned using the Clustawl program provided by the EMBL-EBI database and search engine. Accession numbers for the sequences were as follows: At1g29110; At1g20980; At2g34080; At3g49340; At2g27420. The putative ctVSD is printed in green.
(C) Cysteine proteases bearing a putative N-terminal ssVSD were aligned using the Clustawl program provided by the EMBL-EBI database and search engine. Accession numbers for the sequences were as follows: At2g21430; At4g39090; At4g16190. The putative ssVSD are printed in red.
Blast search was performed using the TIGR database of the Arabidopsis thaliana genome project.
Asterisks are identical amino acids, double points are conserved exchanges, single points are homologous amino acids.

Figure 4 a+b (.jpg)

Figure 4 c (.jpg)