Telomeres are long non-coding regions found at the ends of eukaryotic linear chromosomes. Although they have traditionally been associated with the protection of linear DNA ends to avoid gene losses during each round of DNA replication, recent studies have demonstrated that the role of these sequences and their adjacent regions go beyond just protecting chromosomal ends. Regions nearby to telomeric sequences have now been identified as having increased variability in the form of duplications and rearrangements that result in new functional abilities and biodiversity. Furthermore, unique fungal telomeric and chromatin structures have now extended clinical capabilities and understanding of pathogenicity levels. In this review, telomere structure, as well as functional implications, will be examined in opportunistic fungal pathogens, including Aspergillus fumigatus, Candida albicans, Candida glabrata, and Pneumocystis jirovecii.

Figure legends to supplemental figures 1-6

Supplemental Figure 2. FACS profiles showing DNA content of the synchronized HeLaII at the indicated time points. These correspond to the experiment shown in 3C and 3D.

Supplemental Figure 4. A. IP-TRAP assays showing the precipitation of telomerase activity with antihTERT or anti-FEN1 antibodies, as well as the assays for OBFC1, p68 (POLA2) and POLα (p180). Both p68 and p180 are positive in this assay, whereas OBFC1 was negative. The arrow points to the internal PCR control in the reaction (telomeraseindependent). The asterix indicates a non-specific band. B. IP-TRAP assay performed on HelaII extract (left lane), and on immunoprecipitated material with p180 rabbit serum (S). Controls include RNase treated lysates, as well as beads only and preimmune (PI) controls as indicated.

Supplemental Figure 5. A. Western blot for HeLaII expressing stable p68 shRNA B. In-gel hybridization for HeLaII treated with shRNA for p68 after ten passages. B. Overhang intensity for shRNA samples, processed as described in Fig.5B. C. Quantitation of the overhangs detected as in panel B, with values from three independent experiments. The p-value (two-tailed t-test) is shown for sh644.

Supplemental Figure 6. A. Western blot for HeLaII transfected with the siRNAs indicated on top. The analysis was performed 48 hours post-transfection. B. Telomeric overhang analysis in HelaII depleted for 180 with the indicated siRNAs, using the DSN (double strandspecific nuclease) technique. Briefly, four micrograms of genomic DNA were digested with 0.8 U of double strand-specific nuclease (DSN) as per manufacturer instructions, ran on a 6% PAGE, transferred to a membrane, and hybridized to a C-rich (CCCTAA) probe. Undigested and ExoI controls were ran alongside, as labeled below the gel. See materials and methods, and (32) for further details. C. Quantitation of the overhangs detected in panel B, with values from three independent experiments. The two-tailed Student t test for the two significant values (<0.05) are shown.