DOI: 10.1126/science.284.5413.496. SPA1, a WD-Repeat Protein Specific to Phytochrome A Signal Transduction

SPA1, a WD-Repeat Protein Specific to Phytochrome A Signal Transduction

10.1126/science.284.5413.496

Crossref journal-article

American Association for the Advancement of Science (AAAS)

Science (221)

Abstract

The five members of the phytochrome photoreceptor family of Arabidopsis thaliana control morphogenesis differentially in response to light. Genetic analysis has identified a signaling pathway that is specifically activated by phytochrome A. A component in this pathway, SPA1 (for “suppressor of phyA-105”), functions in repression of photomorphogenesis and is required for normal photosensory specificity of phytochrome A. Molecular cloning of the SPA1 gene indicates that SPA1 is a WD (tryptophan–aspartic acid)-repeat protein that also shares sequence similarity with protein kinases. SPA1 can localize to the nucleus, suggesting a possible function in phytochrome A–specific regulation of gene expression.

Authors 3

Ute Hoecker (first)
James M. Tepperman (additional)
Peter H. Quail (additional)

References 42 Referenced 218

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We used a genomic fragment containing the SPA1 gene to hybridize a cDNA library that was made from mRNA isolated from dark-grown Arabidopsis seedlings that is seedlings grown continuously in complete darkness from germination onward (22). We isolated several cDNA clones the longest of which is 3580 bp in size. This cDNA clone contains three in-frame stop codons preceding the putative ATG start codon. The size of this cDNA is in agreement with a single transcript ∼3.6 kb in length detected in a Northern (RNA) hybridization.
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Single-letter abbreviations for the amino acid residues are as follows: A Ala; C Cys; D Asp; E Glu; F Phe; G Gly; H His; I Ile; K Lys; L Leu; M Met; N Asn; P Pro; Q Gln; R Arg; S Ser; T Thr; V Val; W Trp; and Y Tyr. X indicates any residue.
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We amplified fragments containing candidate genes by polymerase chain reaction (PCR) from genomic DNA isolated from spa1 mutant and wild-type plants. These fragments were sequenced using an ABI373 automated sequencer (Perkin-Elmer). Base-pair changes between spa1 mutant and wild-type plants were found in only one of the candidate genes.
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The NIa coding region in the plasmid pRTL2-GUS/NIaΔBam (23) was replaced by an expanded multiple cloning site: Bgl II–Cla I–Not I–Sal I–BamH I–Xho I. The SPA1 cDNA clone was amplified with primers to create a Cla I site at the ATG and a Sal I site at the TAG digested with these enzymes and inserted into the modified pRTL2-GUS/NIaΔBam plasmid to create GUS:SPA1. This plasmid was then digested with Bgl II to remove a fragment containing the first 1773 bp of the SPA1 coding region and religated to create GUS:SPA1ΔN591.
Total RNA was extracted from 3-day-old seedlings that had been transferred to light for the indicated time using the Qiagen RNeasy Plant Miniprep kit. We separated 5 μg of total RNA on a Mops-RNA gel containing 6.7% formaldehyde and subsequently transferred the RNA to MSI Nylon membrane. The membranes were hybridized with a random prime-labeled PCR product containing the full-length SPA1 cDNA and washed with a final wash with 0.2× saline sodium citrate and 0.1% SDS at 65°C. Membranes were subsequently hybridized with a pea 18 S rRNA probe.
We thank Y. Kang and S. Moran for excellent technical assistance M. Hudson for critical reading of the manuscript D. Aubert for providing aliquots of a binary cosmid library and the Arabidopsis Biological Resource Center in Ohio for providing seeds a cDNA library and clones. Supported by NIH grant GM-47475 and the U.S. Department of Agriculture Current Research Information Service number 5335-21000-006-00D.

Dates

Type	When
Created	23 years, 1 month ago (July 27, 2002, 5:40 a.m.)
Deposited	1 year, 7 months ago (Jan. 13, 2024, 4:31 a.m.)
Indexed	2 days, 5 hours ago (Aug. 27, 2025, 11:53 a.m.)
Issued	26 years, 4 months ago (April 16, 1999)
Published	26 years, 4 months ago (April 16, 1999)
Published Print	26 years, 4 months ago (April 16, 1999)

Funders 0

None

BibTeX

@article{Hoecker_1999, title={SPA1, a WD-Repeat Protein Specific to Phytochrome A Signal Transduction}, volume={284}, ISSN={1095-9203}, url={http://dx.doi.org/10.1126/science.284.5413.496}, DOI={10.1126/science.284.5413.496}, number={5413}, journal={Science}, publisher={American Association for the Advancement of Science (AAAS)}, author={Hoecker, Ute and Tepperman, James M. and Quail, Peter H.}, year={1999}, month=apr, pages={496–499} }

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      "DOI": "10.1016/S0014-5793(97)01558-5"
    },
    {
      "key": "e_1_3_1_27_2",
      "unstructured": "We amplified fragments containing candidate genes by polymerase chain reaction (PCR) from genomic DNA isolated from spa1 mutant and wild-type plants. These fragments were sequenced using an ABI373 automated sequencer (Perkin-Elmer). Base-pair changes between spa1 mutant and wild-type plants were found in only one of the candidate genes."
    },
    {
      "key": "e_1_3_1_28_2",
      "doi-asserted-by": "crossref",
      "first-page": "42",
      "DOI": "10.1126/science.3291115",
      "volume": "241",
      "author": "Hanks S. K.",
      "year": "1991",
      "unstructured": "Hanks S. K., Quinn A. M., Hunter T. H., Science 241, 42 (1991).",
      "journal-title": "Science"
    },
    {
      "key": "e_1_3_1_29_2",
      "unstructured": "The NIa coding region in the plasmid pRTL2-GUS/NIa\u0394Bam (23) was replaced by an expanded multiple cloning site: Bgl II\u2013Cla I\u2013Not I\u2013Sal I\u2013BamH I\u2013Xho I. The SPA1 cDNA clone was amplified with primers to create a Cla I site at the ATG and a Sal I site at the TAG digested with these enzymes and inserted into the modified pRTL2-GUS/NIa\u0394Bam plasmid to create GUS:SPA1. This plasmid was then digested with Bgl II to remove a fragment containing the first 1773 bp of the SPA1 coding region and religated to create GUS:SPA1\u0394N591."
    },
    {
      "key": "e_1_3_1_30_2",
      "unstructured": "Total RNA was extracted from 3-day-old seedlings that had been transferred to light for the indicated time using the Qiagen RNeasy Plant Miniprep kit. We separated 5 \u03bcg of total RNA on a Mops-RNA gel containing 6.7% formaldehyde and subsequently transferred the RNA to MSI Nylon membrane. The membranes were hybridized with a random prime-labeled PCR product containing the full-length SPA1 cDNA and washed with a final wash with 0.2\u00d7 saline sodium citrate and 0.1% SDS at 65\u00b0C. Membranes were subsequently hybridized with a pea 18 S rRNA probe."
    },
    {
      "key": "e_1_3_1_31_2",
      "unstructured": "We thank Y. Kang and S. Moran for excellent technical assistance M. Hudson for critical reading of the manuscript D. Aubert for providing aliquots of a binary cosmid library and the Arabidopsis Biological Resource Center in Ohio for providing seeds a cDNA library and clones. Supported by NIH grant GM-47475 and the U.S. Department of Agriculture Current Research Information Service number 5335-21000-006-00D."
    }
  ],
  "container-title": "Science",
  "original-title": [],
  "language": "en",
  "link": [
    {
      "URL": "https://www.science.org/doi/pdf/10.1126/science.284.5413.496",
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      "content-version": "vor",
      "intended-application": "similarity-checking"
    }
  ],
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    ],
    "date-time": "2024-01-13T09:31:41Z",
    "timestamp": 1705138301000
  },
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  "resource": {
    "primary": {
      "URL": "https://www.science.org/doi/10.1126/science.284.5413.496"
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  },
  "subtitle": [],
  "short-title": [],
  "issued": {
    "date-parts": [
      [
        1999,
        4,
        16
      ]
    ]
  },
  "references-count": 42,
  "journal-issue": {
    "issue": "5413",
    "published-print": {
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        ]
      ]
    }
  },
  "alternative-id": [
    "10.1126/science.284.5413.496"
  ],
  "URL": "http://dx.doi.org/10.1126/science.284.5413.496",
  "relation": {},
  "ISSN": [
    "0036-8075",
    "1095-9203"
  ],
  "subject": [],
  "container-title-short": "Science",
  "published": {
    "date-parts": [
      [
        1999,
        4,
        16
      ]
    ]
  }
}