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By Emily Keator and Phoebe Parrish

By Emily Keator and Phoebe Parrish. : Part. 04-17-14. What Does Iron Do? . Highly reactive—involved in many redox reactions Involved in photosynthesis & cellular respiration as part of the ETC Toxic in high amounts. Uptake Genes. AHA Extrudes H + ions FRO2 Reduces Fe 3+ → Fe 2+

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By Emily Keator and Phoebe Parrish

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  1. By Emily Keator and Phoebe Parrish : Part 04-17-14

  2. What Does Iron Do? • Highly reactive—involved in many redox reactions • Involved in photosynthesis & cellular respiration as part of the ETC • Toxic in high amounts

  3. Uptake Genes • AHA • Extrudes H+ ions • FRO2 • Reduces Fe3+ → Fe2+ • IRT1 • Transports Fe2+ into cell (regulated by ubiquitin) • Also involved in uptake of other heavy metal cations

  4. Transport Genes • YSL1&2 • Influx (phloem) • Transport Fe-NA complexes • FRD3 (MATE efflux family proteins) • Efflux (xylem) • Transport Fe-citrate complexes; provide shoots with usable forms of Fe

  5. Storage Genes • Ferritin • Complexes Fe2+ molecules together • Stores readily-usable form of Fe in vacuoles & amyloplasts • PIC1 • Transports Fe into chloroplasts • IREG1 • Moves Fe into vacuole

  6. Regulatory Genes • bHLH38,39,100&101 • Regulate IRT1 and FRO2 genes • FIT1 • Forms heterodimers with bHLH → control transcription of Fe uptake genes • Both transcribed when Fe deficiency is sensed in the roots

  7. QTLs

  8. BLAST Results

  9. QTL * = confirmed w/ IGB Questionable presence on IGB 0 -10,000,000 QTL C01: 1,923,307 IRT1: 3,007,445* AHA1,2,9: 4,691,639* C01 C03 FRD3: 12,416,349* QTL C03: 13,636,922 FIT1: 6,411,902* FIT1: 16,455,349* AHA1,2,9: 19,835,760* YSL2: 11,431,126* YSL1: 11,431,687 +10,000,000 +10,000,000

  10. QTL * = confirmed w/ IGB Questionable presence on IGB -10,000,000 -10,000,000 FPN1/IREG1:34,728,143 FIT1: 39,453,920 FER1:30,370,173* bHLH38&39: 30,863,851* IRT1: 41,537,794* bHLH100&101: 30,878,266, 30,890,636* FER1: 42,532,200* AHA1,2,9: 32,394,505* C05 C08 QTL C05: 43,302,017 QTL C08: 33,379,515 FRD3: 43,624,447* AHA1,2,9: 43,882,387* IRT1:39,033,254* FRO1,2: 41,520,823* +10,000,000 +10,000,000

  11. QTL * = confirmed w/ IGB Questionable presence on IGB -10,000,000 0 FRO1: 137,473* YSL2: 36,138,179* AHA1, AHA2: 514,992* YSL1,2: 1,879,450* AHA1,2,9:40,597,697* YSL1: 40,697,628 QTL C09-1: 3,558,943 C09 C09 QTL C09-2: 45,002,150 PIC1: 7,315,088* IRT1: 49,050,708* AHA9: 8,682,603* +10,000,000 +10,000,000

  12. IGB Results

  13. -1,000,000 QTL Confirmed BLAST result Genes found only on IGB ^ = gene identified through BLAST/TAIR method C01 QTL C01: 1,923,307 2,114,850: Heavy metal transport/detoxification superfamily protein^ 3,007,445 (ZIP9)^ +1,000,000

  14. Confirmed BLAST result Genes found only on IGB • ZIP9 (homolog of IRT3) • Location: nucleus, plasmodesma • Expression: petal differentiation & expansion stage • Function: transports zinc and other cations; contains zinc/iron permease domain • Heavy metal transport/detoxification superfamily protein • Location: nucleus, plasmodesma • Expression: petal differentiation and expansion stage • Function: metal ion transport and binding

  15. -1,200,000 QTL Confirmed BLAST result Genes found only on IGB ^ = gene identified through BLAST/TAIR method 12,415,632: MATE efflux family protein (FRD3)^ 12,418,215: Multidrug resistance protein^ C03 QTL C03: 13,636,922 +1,200,000

  16. Confirmed BLAST result Genes found only on IGB • FRD3 (MATE efflux family protein) • Location: plasma membrane • Expression: petal differentiation & expansion stage • Function: iron-citrate transport • MdtK (multidrug resistance protein) • Location: chloroplast & plasma membrane • Expression: petal differentiation & expansion stage • Function: ion transporter/antiporter activity

  17. 41,537,794: zinc transporter (IRT1)^ -1,000,000 QTL Confirmed BLAST result Genes found only on IGB ^ = gene identified through BLAST/TAIR method 42,532,200: ferritin (FER1)^ 42,731,745: Heavy metal transport/detoxification superfamily protein^ C05 QTL C05: 43,302,017 43,882,387: calcium transporting ATPase^ 43,536,616: MATE efflux family protein (FRD3)^ 43,624,447: MATE efflux family protein (FRD3)^ +1,000,000

  18. Confirmed BLAST result Genes found only on IGB • IRT1 • Location: membrane • Expression: petal differentiation & expansion stage • Function: cellular response to iron ion starvation, iron transport, etc. • FER1 • Location: chloroplast • Expression: petal differentiation & expansion • Function: iron ion transport and homeostasis, ferric iron binding • Heavy metal transport/detoxification (homolog of ATX1) • Location: plasma membrane • Expression: pollen, petal, petiole • Function: metal ion transport & binding • FRD3 (MATE efflux family protein) • Location: plasma membrane • Expression: roots, flower, petiole • Function: iron ion and citrate transport & homeostasis

  19. -1,000,000 QTL Confirmed BLAST result Genes found only on IGB ^ = gene identified through BLAST/TAIR method 32,394,505: H+-ATPase (AHA) C08 QTL C08: 33,379,515 33,828,404: Ferrochelatase +1,000,000

  20. Confirmed BLAST result Genes found only on IGB • AHA (H+-ATPase) • Location: plasma membrane, Golgi apparatus, vacuolar membrane • Expression: petal differentiation & expansion • Function: ion transport • Ferrochelatase • Location: chloroplast • Expression: petal differentiation & expansion • Function: terminal enzyme of heme biosynthesis

  21. -1,000,000 QTL Confirmed BLAST result Genes found only on IGB ^ = gene identified through BLAST/TAIR method 3,501,200: Ferrochelatase^ C09 QTL C09-1: 3,558,943 +1,000,000

  22. QTL Confirmed BLAST result Genes found only on IGB ^ = gene identified through BLAST/TAIR method -1,000,000 C09 QTL C09-2: 45,002,150 45,568,877: MATE efflux family protein 45,927,322: heavy metal transport/detoxification superfamily protein +1,000,000

  23. Confirmed BLAST result Genes found only on IGB • MATE efflux family protein • Location: (?) • Expressed in: (?) • Function: transport • Heavy metal transport/detoxification superfamily protein • Location: extracellular space (?) • Expression: petal differentiation & expansion • Function: exact function unknown

  24. Conclusions • IGB contained almost all genes found on BLAST • Used IGB to confirm genes found on BLAST (+/- 1,000,000 bp) • IGB yielded several new candidates (more speculative) • Overall: a number of good candidate genes, close to QTL and expressed in relevant tissues

  25. References Barberon M et al. Monoubiquitin-dependent endocytosis of the iron-regulated transporter 1 (IRT1) transporter controls iron uptake in plants. PNAS, 2011. 108(32):E450-E458. Briat, JF et. al. Regulation of plant ferritin synthesis: how and why. Cellular and Molecular Life Sciences, 1999. 56:155-166. Grotz, N et. al. Molecular aspects of Cu, Fe, and Zn homeostasis in plants. Biochemica et Biophysica Acta, 2006. 1763:595-608. Harper JF et al. The Arabidopsis thaliana plasma membrane H+-ATPase multigene family. The Journal of Biological Chemistry, 1990. 265(23):13601-13608. Hell, R et. al. Iron uptake, trafficking, and homeostasis in plants. Planta, 2003. 216:541-551. Kim, S et. al. Mining iron: iron uptake and transport in plants. FEBS Letters, 2007. 581:2273-2280. Kobayashi T, Nishizawa NK. Iron uptake, translocation, and regulation in higher plants. Annual Review of Plant Biology, 2012. 63:131-52. Matsuoka, K et. al. Gibberellin-induced expression of Fe uptake-related genes in Arabidopsis. Plant Cell Physiol., 2014. 55(1):87-98. Robinson, NJ et. al. A ferric-chelate reductase for iron uptake from soil. Nature, 1999. 397:694-697. Roschzttardtz, H et. al. New insights into Fe localization in plant tissues. Frontiers In Plant Science, 2013. 4:1-11. Seo, PJ et. al. A golgi-localized MATE transporter mediates iron homeostasis under osmotic stress in Arabidopsis. BIochemical Journal, 2012. 442:551-561. Sussman MR. Molecular analysis of proteins in the plant plasma membrane. Annual Review of Plant Physiology and Plant Molecular Biology, 1994. 45:211-34. Vert, G et. al. IRT1, an Arabidopsis transporter essential for iron uptake from the soil and for plant growth. The Plant Cell, 2002. 14:1223-1233.

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