Supplementary Fig. 1. Infection of three C. elegans strains used for spatially restricted enzymatic tagging. Animals infected with N.
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1 Supplementary Fig. 1. Infection of three C. elegans strains used for spatially restricted enzymatic tagging. Animals infected with N. parisii stained with a FISH probe (red) specific for Nematocida rrna.
2 Supplementary Fig. 2. Biotin labeling of infected C. elegans displays specificity for the host intestinal tissue. Dissected intestines stained with anti-gfp antibody (green), with DAPI for DNA (blue) and with Alexa Fluor 568 streptavidin to label biotin. Left, GFP only. Middle, GFP- APX-NES Right, GFP-APX-NLS. CEN, C. elegans intestinal nuclei. Labeled arrows indicate representative C. elegans nuclei, which are large blue circles of DNA. NPN, N. parisii nuclei. Labeled arrows indicate representative N. parisii nuclei, which are small blue circles of DNA.
3 Supplementary Fig. 3. The number of C. elegans and microsporidia proteins identified through spatially restricted enzymatic tagging. C. elegans (blue) or Nematocida (orange) proteins identified from animals infected with either N. parisii or N. sp. 1. Numbers represent the total proteins with at least 1 peptide identification from either GFP, GFP-APX-NES, or GFP-APX- NLS. Supplementary Fig. 4. Properties of identified C. elegans intestinal proteins. A. Comparison of identified C. elegans proteins specific to the nucleus (118 proteins) to those specific to the cytoplasm (114 proteins). GO term analysis for the compartments of the nucleus and the cytoplasm was performed using PANTHER 55. B. Properties of 891 C. elegans proteins identified in the intestine with spatially restricted enzymatic tagging, compared to the C. elegans genome. TM, transmembrane. SP, signal peptide.
4 Supplementary Fig. 5. N. parisii proteins in the no APX sample are not enriched in the same properties as host-exposed proteins. A. Comparison of mrna expression levels of the 72 identified host-exposed proteins (orange), 392 proteins identified in the No APX sample (green), and the rest of N. parisii genome (blue). Expression data are from animals infected for 30 hours at 25 C 49. B. Comparison of the 72 identified host-exposed proteins (orange), 392 proteins identified in the No APX sample (green), and the genome (blue). Left, categorization by properties enriched in host-exposed proteins. TM, transmembrane. SP, signal peptide. Right, categorization by conservation class. The percentage of the N. parisii genome, no APX proteins, and hostexposed proteins in each category are shown.
5 Supplementary Fig. 6. Phylogenetic tree of 21 microsporidia species. Phylogenetic tree of 21 microsporidia species and R. allomycis as an outgroup. Tree was inferred using RAxML using the PROTGAMMALG model and 1000 bootstrap replicates. Bootstrap support is indicated next to each node. Scale bar indicates changes per site. Strains are in parentheses. Tree was created using FigTree (
6 Supplementary Fig. 7. Properties of N. sp. 1 host-exposed proteins. Identified host-exposed proteins (orange) were compared to all proteins in the genome (blue). SP/TM, signal peptide or transmembrane domain. The percentage of the N. sp. 1 genome and the N. sp. 1 host-exposed proteins in each category are shown. Enrichment p-values (one-side Fisher s exact test) are listed below each category. Supplementary Fig. 8. Properties of Spraguea lophii proteins identified from germinated spores. Proteins identified from germinated S. lophii spores (orange) were compared to all proteins in the genome (blue). SP, signal peptide. The percentage of the S. lophii genome and the S. lophii secreted proteins in each category are shown. Enrichment p-values (one-side Fisher s exact test) are listed below each category. Proteins identified from germinated S. lophii spores is taken from Campbell et al.
7 Supplementary Fig. 9. Evidence of duplication of large gene family proteins. A. Percentage of large gene family proteins in N. parisii at chromosomal ends (See methods). B. Percentage of large gene family proteins in N. parisii that are adjacent to each other (See methods).
8 Supplementary Fig. 10. Pfam domains in predicted host-exposed proteins that are not part of large gene families. Pfam domains with at least 8 occurrences in predicted host-exposed proteins that are not members of large gene families are shown.
9 Supplementary Table 1. Strains used in this study. Tissue (promoter) Protein Subcellular location (localization tag) Strain nomenclature intestine (spp-5) GFP none ERT383 jysi11 [pet536(spp-5p::gfp::unc-54 ; unc-119 (+)]II; unc-119(ed3) III intestine (spp-5) GFP-APX cytoplasm (NES) ERT385 jysi13 [pet532(spp-5p::gfp_apx_nes::unc-54 ; unc-119 (+)]II; unc-119(ed3) III intestine (spp-5) GFP-APX nucleus (NLS) ERT384 jysi12 [pet522(spp-5p::gfp_apx_nls::unc-54 ; unc-119 (+)]II; unc-119(ed3) III Supplementary Table 2. Predicted signal peptidase-processed N-terminal peptides. Protein Peptide[a] GFP1[b] GFP2[b] GFP3[b] NES1[b] NES2[b] NES3[b] NLS1[b] NLS2[b] NLS3[b] NEPG_ S]SFVLGNAANHASTPTR.S NEPG_ T]SAINHNPEVDAPQEM*SK.E NEPG_ KAESADPSTTLANAGSDK.G NEPG_ FIEQYKPNSPIIVIK.E NEPG_ VNYIQETVFSPEFSK.G NEPG_ VIQQQYSNVEIR.K NEPG_ KFTFENIEEVQR.T NEPG_ LKPLVFNLPSDSPSLR.E NERG_ INYIQESIFSPEFSK.G NERG_ S]MGIHSIEK.V [a] "-." is the N-terminal, "]" is N-terminal acetylation, * is oxidized Methionine and "X.X" is the trypsinization site. [b] The number of peptide counts in each sample. Supplementary Table 3. Summary of genome assembly statistics. Species Strain (assembly) Total sequence length Total assembly gap length Number Contig Contig of contigs N50 L50 Number of scaffold s Scaffold N50 Scaffold L50 Proteins % Orthogroups conserved Spraguea lophii 42_110 (Sprlop1.0) N/A N/A N/A Nosema bombycis CQ1 (NosBomCQ1_v1.0) Mitosporidium daphniae (UGP1.0) N/A N/A N/A Nematocida displodere JUm2807 (Nema_displodere_ERTm0_V1) Nematocida ironsii ERTm5 (Nema_ironsii_ERTm5_V1) Nematocida sp. 1 ERTm2 (Nema_parisii_ERTm2_V1) Nematocida sp. 1 ERTm6(Nema_sp_1_ERTm6_V2) 4,276,041 47, ##### , Nematocida parisii ERTm1(Nema_parisii_ERTm1_V3) 4,071,346 42, ##### , Nematocida parisii ERTm3 (Nema_parisii_ERTm3_V1) Ordospora colligata OC4 (ASM80326v1) N/A N/A N/A Enterocytozoon bieneusi H348 (ASM20948v1) Encephalitozoon hellem ATCC (ASM27781v3) N/A N/A N/A Trachipleistophora hominis (ASM31613v1) Encephalitozoon cuniculi GB-M1 (ASM9122v1) N/A N/A N/A Encephalitozoon intestinalis ATCC (ASM14646v1) N/A N/A N/A Encephalitozoon romaleae SJ-2008 (ASM28003v2) N/A N/A N/A Vittaforma corneae ATCC (Vitt_corn_V1) Vavraia culicis subsp. floridensis (Vavr_culi_floridensis_V1) Edhazardia aedis USNM (Edha_aedis_V4b) N/A N/A N/A Anncaliia algerae PRA339 (Annc_alge_insect_USDA_JJB_V2 ) Nosema apis BRL 01 (NapisBRLv01) Nosema ceranae PA (ASM98816v1) N/A N/A N/A Pseudoloma neurophilia MK1 (ASM143216v1) 5,248, N/A N/A N/A
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