Rapid Senescence in Sockeye Salmon
       (Oncorhynchus nerka)
Rapid Senescence
• Pacific salmon are semelparous and die of
  senescence soon after beginning reproductive
  behavior

• ...
Physical Signs of Senescence


 Frayed fins




                      Loss of coloration
Research Question:

WHAT CAUSES RAPID SENESCENCE
IN SOCKEYE SALMON?
Study Site in Southwest Alaska:
         Hansen Creek
               Small tributary of Lake Aleknagik

               Nea...
Study Site in Southwest Alaska:
             Iliamna Lake Ponds
Tributary of Lake Iliamna, the
largest lake in Alaska (2,6...
Salmon Entering Hansen Creek
              Up to 40% of individuals may strand
              at stream mouth in a given ye...
Laboratory Procedures
• RNA extraction and quantification of genes of
  interest (brain tissues)
  – Adrenocorticotropic h...
Why did I look at ACTH & CRH?

                         Both are genes involved in
                         the cortisol p...
Why heat shock proteins?
• Heat shock proteins = stress proteins
  – Chaperones: move other proteins around cells
  – Help...
Gene Expression Analysis

                 Designed primers for ACTH,
                 CRH, and HSC71

                 Qu...
No significant differences
                                          or major trends
                      25
Relative Exp...
Explanations for high variation?
• Factors I explored for each gene of interest:
  – Body size
  – Sex of individuals
  – ...
CRH levels lower in
                                 slower-senescing individuals
                      35.0

            ...
Western Blot Protein Analysis




Western Blot using antibody for HSP70

Quantified total protein from each sample and HSP...
Increased concentrations of HSP70
                              in senescent individuals
                 8000

          ...
Summary of Findings
• High CRH levels indicate or result in more
  rapid senescence

• HSP70 at higher levels in senescent...
Prospects for Future Research
• Follow up on CRH findings, directly measure
  cortisol levels

• Look at other genes that ...
Jensen FISH495
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Presentation on salmon senescence as part of FISH495 course

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  • Ran proteins on SDS-PAGE to separate proteins by molecular weightProteins transferred to a nylon membrane
  • Jensen FISH495

    1. 1. Rapid Senescence in Sockeye Salmon (Oncorhynchus nerka)
    2. 2. Rapid Senescence • Pacific salmon are semelparous and die of senescence soon after beginning reproductive behavior • Individuals go from peak health to death in a matter of days to weeks – Rates vary within populations and among populations • Loss of aggression, loss of strength and swimming ability, and physical decline
    3. 3. Physical Signs of Senescence Frayed fins Loss of coloration
    4. 4. Research Question: WHAT CAUSES RAPID SENESCENCE IN SOCKEYE SALMON?
    5. 5. Study Site in Southwest Alaska: Hansen Creek Small tributary of Lake Aleknagik Nearly uniform top to bottom: 4m wide & 10cm deep riffles Annual spawning population of 3,000 – 20,000 sockeye salmon Stream life annually averages below 10 days (usually 7-14)
    6. 6. Study Site in Southwest Alaska: Iliamna Lake Ponds Tributary of Lake Iliamna, the largest lake in Alaska (2,600 km2) Small, variable population size (~500) Larger individuals and longer reproductive lifespan (19.4 days in 2008)
    7. 7. Salmon Entering Hansen Creek Up to 40% of individuals may strand at stream mouth in a given year Sampled for a “pre-senescent” baseline for gene expression
    8. 8. Laboratory Procedures • RNA extraction and quantification of genes of interest (brain tissues) – Adrenocorticotropic hormone (ACTH) – Corticotropin-releasing hormone (CRH) – Heat shock protein (HSC71) • Protein extraction and quantification (liver tissues) – Heat shock protein: HSP70
    9. 9. Why did I look at ACTH & CRH? Both are genes involved in the cortisol production and regulation pathway Cortisol = stress hormone •Suppresses immune system •Inhibits glucose uptake •Stimulates protein degredation Anterior Kidney
    10. 10. Why heat shock proteins? • Heat shock proteins = stress proteins – Chaperones: move other proteins around cells – Help with protein-folding & identify incorrectly- folded proteins – Prepare old proteins to be degraded by cell • Predicted to be associated with senescence
    11. 11. Gene Expression Analysis Designed primers for ACTH, CRH, and HSC71 Quantified with Real-time PCR Corrected for total RNA used by comparing 18s expression levels
    12. 12. No significant differences or major trends 25 Relative Expression 20 CRH 3 ACTH Relative Expression 15 10 2 5 0 1 Pre-Senescent Hansen Ponds Senescent Senescent 0 40 Pre-Senesncent Hansen Ponds Relative Expression 35 HSC71 Senescent Senescent 30 25 20 15 10 • Large Variation & 5 0 Low Sample Sizes Pre-Senescent Hansen Ponds Senescent Senescent
    13. 13. Explanations for high variation? • Factors I explored for each gene of interest: – Body size – Sex of individuals – Entry date or date of senescence – Stream life
    14. 14. CRH levels lower in slower-senescing individuals 35.0 30.0 R² = 0.903 Relative Expression P = 0.0036 25.0 20.0 15.0 10.0 5.0 0.0 3 4 5 6 7 8 9 In-stream Life (Days)
    15. 15. Western Blot Protein Analysis Western Blot using antibody for HSP70 Quantified total protein from each sample and HSP70 proportion
    16. 16. Increased concentrations of HSP70 in senescent individuals 8000 7000 6000 Band Intensity 5000 4000 3000 2000 1000 0 Pre-Senescent Hansen Senescent Ponds Senescent Sampling Group
    17. 17. Summary of Findings • High CRH levels indicate or result in more rapid senescence • HSP70 at higher levels in senescent individuals – Slightly higher in the more rapidly senescing Hansen Creek population than in Iliamna Ponds
    18. 18. Prospects for Future Research • Follow up on CRH findings, directly measure cortisol levels • Look at other genes that may be involved in rapid senescence

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