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Mutations in Paramecium CaM
Indicate Functional Differences b/w
the C-Terminal & N- Terminal Lobes
In Vivo
JA Kink, ME Maley, RR Preston, KY
Ling, MA Wallen-Friedman, Y Saimi
& C Kung
Ashim Malhotra
October 27, 2003
Lay Out
 Background & Introduction
 CaM Structure: Rasmol
 Experimentations
 Conclusions
 Philosophy & Trends
Calmodulin
 148 amino acid long protein with 4 calcium ion
binding pockets-I, II, III, IV.
 III & IV in C-terminal loop & I & II in N-terminal
loop.
 Mediates many Ca signaling pathways.
 Ca ion binding causes a “closed” conformation with
hydrophobic domains inaccessible to targets.
Calmodulin Structure
www.angelfire.com/d20/bio_lab
Paramecium Movement
http://micro.magnet.fsu.edu/moviegallery/ponds
cum/protozoa/paramecium/
Immunoblots of Mutant & wt CaMs
Nucleotide Sequence of P. tetraurelia
CaM gene
Analysis of Paramecium CaM gene
Northern Blot
Southern Blot
Amino Acid Sequence of wt &
Mutant CaM genes
Effect of CaM Mutations on Na+
Behavior & Ca2+ Currents
Ca2+ -Dependent Na+ Current & Ca2+ - Dependent
K+ Current of wt, Cam3 & Cam12 Strains of P.
teraurelia
CaM Electrophoretic Mobilities Cosegregate w/
Behavioral Phenotypes among Autogamous F2
Segregants
Conclusions
 CaM mutation may not be lethal, instead it
may cause behavioral/functional mutations.
 In P. tetraurelia, behavior may be directly
influenced by genetic mutations- the genetic
dissection method for studying mutational
effects.
 The “over-reactor” mutants carry mutations
in amino terminal half of CaM gene.
Conclusions (contd…)
 The “under-reactor” mutants carry mutations
in C-terminal half of the gene.
 The “under-reactors” show low inward Ca2+ Dependent Na+ Currents.
 The “over-reactors” show low Ca2+ Dependent K+ Currents.
The Philosophy
 Functional or behavioral responses have a genetic
origin ~ anthropological approaches for studying
human development, particularly of intelligence.
 A molecular phenomenology sited as causative
link in an evolutionary process.
Thank You