Table 1 Frequencies of illegitimate mating in tetracycline- regulatable promoter conditional alleles grown in the presence and absence of doxycycline Ddc2 foci and high levels of illegitimate mating (Figure 2C). These strains were subjected to a quantitative illegitimate mating assay (Table 1). In the presence of doxycycline, all of these strains exhibited significantly elevated levels of ille- gitimate mating relative to the wild-type strain. Increases in illegitimate mating ranged from <2-fold wild type (CSE1) to 62-fold wild type (MCM7). Previous studies of GAL promoter- regulated conditional alleles of DNA polymerases a and 8 found increases of 200-fold and 50-fold, respectively (Lemoine et al 2005, 2008). Although DNA polymerases a and 8 were not assayed in our screens, we identified a role for DNA polymerase e (POL2 and DPB11) in suppressing illegiti- mate mating. Additionally, we found that disrupting a wide range of replication functions (CDC45, MCM4, MCM5, MCM7, DPB11, POL2, POL30, RFC2, and RFC5) caused increased ille- gitimate mating. DNA2, which functions in Okazaki fragment processing (Budd et al 2000; Lee et al. 2000) and in DNA repair (Zhu et al. 2008) resulted in increased illegitimate mat- ing, as did repression of the DNA repair genes NSE1 (Santa Maria et al. 2007; Pebernard et al. 2008) and UBC9 (Branzei et al. 2006). Genes with functions outside of DNA replication and repair were also identified. CSE1 is responsible for nuclear shuttling of the nuclear transporter importin a (Hood and Silver 1998; Kunzler and Hurt 1998; Solsbacher et al 1998), and roles for CSE1 in DNA replication (Yu et al. 2006) and in chromosome segregation (Xiao et al. 1993), likely reflecting effects on importin a cargos, have been de- scribed. NUF2 is a kinetochore component and functions in chromosome segregation (Osborne et al. 1994). Depletion of Frequencies of illegitimate mating (10-4) +doxycycline Tet allele -doxycycline Wild type CDC45 [1] 14 [0.6) [5.2 [1.1 (13] 14.1 [4.9 (21 [3.0 (2.6 6.5) [2.6) [1.8 [2.9 [2.9 0.83 (0.20) 0.8 (0.26) 1]1 (221 (6.0) (0.04) (2.7) (1.6) (0.40) (2.5) (3.4) (27) 11 17 1.8 [5.9 12 [45 [15] [62) [48) [22) [331 CSE1 0.5 1.4 4.3 4.7 DNA2 DPB11 0.94 (0.1) (9.8) (0.8) (2.9) (11) (1.1) (0.9) (4.9) (0.03) (0.76) (1.7) (0.4) 9.2 36 MCM4 11 cMS MCM7 NSE1 3.4 12 (3.0) 34 (24) (7.3) (8.9) (21) (2.7) (0.14) (0.64) (11) 4.1 39 17 2.5 18 NUF2 POL2 2.2 [10] [7.4 і3 81 8.3 5.4 POL30 5.9 3.0 RFC2 2.2 RFCS 1.5 2.4 [14] (4.8 11 SPT16 3.9 (2.2) UBC9 2.4 Tet allele strains and wild-type strain were grown in parallel for 24 hr on YPD liquid media containing or lacking 10 ug/ml doxycycline. Strains were mixed with fivefold excess of a MATa tester strain and plated on YPD solid media. After 5 hr, cells were resuspended in water and plated on illegitimate diploid selection media. This assay was repeated two times. Numbers in parentheses represent standard deviations Numbers in brackets represent the frequency normalized to wild type. although the overlap between the two screens, at 29 genes, was not absolute. We focused on the strains with the most robust chromo- some instability phenotype, the 15 strains with both elevated 1225a Tester Strain Tet Allele A thr4 MATa THR X MATa his4 HIS4 Figure 3 Classification of rear- rangement events that lead to il- legitimate mating. (A) Schematic diagram of the three expected CLASS 3 CLASS 2 Chromosome III loss CLASS 1 Gene conversion or mutation Chromosome II right arm loss classes of genetic events result- ing in illegitimate mating. Using (HIS4 HIS4 mata-x THR4 diagnostic selection media, mu- tations in the MAT locus, whole chromosome II loss, and loss of the right arm of chromosome III can be distinguished as class 1, 2, MATa thr4 MATa thr4 his4 his4 MATa thr4 his4 and 3 genetic events, respectively er (Lemoine et al. 2005, 2008). (B) We classified ~200 illegitimate 1.0 В 3.0 0.8 2.5 diploids for each strain. The fre- uencies of the three classes of पा স rearrangements are plotted for the 15 strains with the most ele- vated levels of illegitimate mating (C) Ratios of the three classes of 2.0 0.6 1.5 0.4 1.0 0.2 rearrangements are plotted for the indicated strains. 0.5 0 CLASS 1 CLASS 3 CLASS 3 CLASS 2 CLASS CLASS 2 Tet Alleles Tet Alleles Essential Genome Stability Genes 151 Frequency of I Diploid Classimate 20H 9uds 켜60an Ratios of Illegitimate Diploids wildtype CDC45 CSE1 DNA2 DPB11 MCM4 MCM7 NSE1 NUF2 6280

Table 1 Frequencies of illegitimate mating in tetracycline- regulatable promoter conditional alleles grown in the presence and absence of doxycycline Ddc2 foci and high levels of illegitimate mating (Figure 2C). These strains were subjected to a quantitative illegitimate mating assay (Table 1). In the presence of doxycycline, all of these strains exhibited significantly elevated levels of ille- gitimate mating relative to the wild-type strain. Increases in illegitimate mating ranged from <2-fold wild type (CSE1) to 62-fold wild type (MCM7). Previous studies of GAL promoter- regulated conditional alleles of DNA polymerases a and 8 found increases of 200-fold and 50-fold, respectively (Lemoine et al 2005, 2008). Although DNA polymerases a and 8 were not assayed in our screens, we identified a role for DNA polymerase e (POL2 and DPB11) in suppressing illegiti- mate mating. Additionally, we found that disrupting a wide range of replication functions (CDC45, MCM4, MCM5, MCM7, DPB11, POL2, POL30, RFC2, and RFC5) caused increased ille- gitimate mating. DNA2, which functions in Okazaki fragment processing (Budd et al 2000; Lee et al. 2000) and in DNA repair (Zhu et al. 2008) resulted in increased illegitimate mat- ing, as did repression of the DNA repair genes NSE1 (Santa Maria et al. 2007; Pebernard et al. 2008) and UBC9 (Branzei et al. 2006). Genes with functions outside of DNA replication and repair were also identified. CSE1 is responsible for nuclear shuttling of the nuclear transporter importin a (Hood and Silver 1998; Kunzler and Hurt 1998; Solsbacher et al 1998), and roles for CSE1 in DNA replication (Yu et al. 2006) and in chromosome segregation (Xiao et al. 1993), likely reflecting effects on importin a cargos, have been de- scribed. NUF2 is a kinetochore component and functions in chromosome segregation (Osborne et al. 1994). Depletion of Frequencies of illegitimate mating (10-4) +doxycycline Tet allele -doxycycline Wild type CDC45 [1] 14 [0.6) [5.2 [1.1 (13] 14.1 [4.9 (21 [3.0 (2.6 6.5) [2.6) [1.8 [2.9 [2.9 0.83 (0.20) 0.8 (0.26) 1]1 (221 (6.0) (0.04) (2.7) (1.6) (0.40) (2.5) (3.4) (27) 11 17 1.8 [5.9 12 [45 [15] [62) [48) [22) [331 CSE1 0.5 1.4 4.3 4.7 DNA2 DPB11 0.94 (0.1) (9.8) (0.8) (2.9) (11) (1.1) (0.9) (4.9) (0.03) (0.76) (1.7) (0.4) 9.2 36 MCM4 11 cMS MCM7 NSE1 3.4 12 (3.0) 34 (24) (7.3) (8.9) (21) (2.7) (0.14) (0.64) (11) 4.1 39 17 2.5 18 NUF2 POL2 2.2 [10] [7.4 і3 81 8.3 5.4 POL30 5.9 3.0 RFC2 2.2 RFCS 1.5 2.4 [14] (4.8 11 SPT16 3.9 (2.2) UBC9 2.4 Tet allele strains and wild-type strain were grown in parallel for 24 hr on YPD liquid media containing or lacking 10 ug/ml doxycycline. Strains were mixed with fivefold excess of a MATa tester strain and plated on YPD solid media. After 5 hr, cells were resuspended in water and plated on illegitimate diploid selection media. This assay was repeated two times. Numbers in parentheses represent standard deviations Numbers in brackets represent the frequency normalized to wild type. although the overlap between the two screens, at 29 genes, was not absolute. We focused on the strains with the most robust chromo- some instability phenotype, the 15 strains with both elevated 1225a Tester Strain Tet Allele A thr4 MATa THR X MATa his4 HIS4 Figure 3 Classification of rear- rangement events that lead to il- legitimate mating. (A) Schematic diagram of the three expected CLASS 3 CLASS 2 Chromosome III loss CLASS 1 Gene conversion or mutation Chromosome II right arm loss classes of genetic events result- ing in illegitimate mating. Using (HIS4 HIS4 mata-x THR4 diagnostic selection media, mu- tations in the MAT locus, whole chromosome II loss, and loss of the right arm of chromosome III can be distinguished as class 1, 2, MATa thr4 MATa thr4 his4 his4 MATa thr4 his4 and 3 genetic events, respectively er (Lemoine et al. 2005, 2008). (B) We classified ~200 illegitimate 1.0 В 3.0 0.8 2.5 diploids for each strain. The fre- uencies of the three classes of पा স rearrangements are plotted for the 15 strains with the most ele- vated levels of illegitimate mating (C) Ratios of the three classes of 2.0 0.6 1.5 0.4 1.0 0.2 rearrangements are plotted for the indicated strains. 0.5 0 CLASS 1 CLASS 3 CLASS 3 CLASS 2 CLASS CLASS 2 Tet Alleles Tet Alleles Essential Genome Stability Genes 151 Frequency of I Diploid Classimate 20H 9uds 켜60an Ratios of Illegitimate Diploids wildtype CDC45 CSE1 DNA2 DPB11 MCM4 MCM7 NSE1 NUF2 6280

Human Heredity: Principles and Issues (MindTap Course List)

11th Edition

ISBN:9781305251052

Author:Michael Cummings

Publisher:Michael Cummings

Chapter6: Cytogenetics: Karyotypes And Chromosome Aberrations

Section: Chapter Questions

Problem 14QP: Variations in Chromosome NumberAneuploidy Describe the process of nondisjunction and explain when it...

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