Figure 1

pan-1 gene structure and mRNA isoform expression. (A) mRNA splicing pattern of pan-1 and predicted encoded proteins. RT-PCR data indicate three SL1 trans-splice sites that would generate three distinct mRNA isoforms: pan-1A, pan-1B, and pan-1C. The isoforms generated by the SL1 trans-splice sites are indicated in parentheses. Possible translational start codons for the different isoforms are also indicated. The encoded PAN-1B protein isoform has 11 LRRs (vertical stippled boxes) as identified by the SMART program. An additional 4 LRRs are predicted by LRRscan, as described in Dolan et al. 2007 (yellow boxes). The N and C terminal LRR capping domains (blue colored regions) were identified using the consensus sequence found in Dolan et al., 2007. The PAN-1 N-terminal capping domain (LRR-NT) has the amino sequence CIDIEKGFKEEFNAHKQP VCICADNGIFSTVKGFTIEC. The C-terminal PAN-1 capping domain (LRR-CT1) has the amino acid sequence PWVCVCNDPKEWLPRWLEASEEADVAEGALGCLAIPNC. The signal peptide (‘SP’) is indicated by the red colored region and the transmembrane domain (‘TM’) is indicated by the purple colored region. The cytoplasmic domain (‘CD’) and GFP fusion point (green bar) are also indicated. PAN-1A is not predicted to encode a transmembrane protein and contains fewer LRRs. (B) RT-PCR analysis of pan-1 on total RNA isolated from glp-1(bn2ts) and also wild-type adults subjected to control or pan-1 RNAi (1% agarose gel stained with ethidium bromide). glp-1(bn2ts) larvae raised to adulthood at 16°C form a germline while animals raised at 25°C lack germline tissue. pan-1A and pan-1B are amplified from cDNA generated from both populations while pan-1C is only robustly amplified from cDNA generated from animals raised at 16°C, indicating germline enrichment of pan-1C relative to the soma. All three isoforms are also amplified from cDNA generated from wild-type adult animals. unc-54 is a somatic gene that is expressed in both 16°C and 25°C cDNA preparations.