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in DSB repair became apparent in pachynema upon probing of mutant spermatocyte nuclei with antibodies against molecules involved in various stages of recombination. In \u003e99% of Trip13Gt/Gt chromosome spreads, BLM helicase (Figure 4C and 4D), RAD51/DMC1 (Figure 4E and 4F), γH2AX (Figure 4G and 4H), and TOPBP1 (Figure 4I and 4J) all persisted abnormally on synapsed chromosomes. For RAD51/DMC1, mutant pachytene spermatocytes contained 138 ± 6 foci (compared to 11 ± 3 foci in wild type, most of which were on the XY body), down from 218 ± 13 in zygonema (compared to 220 ± 13 foci in wild type). TOPBP1 is a DNA damage–checkpoint protein involved in ATM protein–dependent activation of ATR protein [20,21]. It binds sites of DSBs and unsynapsed regions of meiotic chromosomes [22,23]. BLM has been reported to colocalize with markers (RPA and MSH4) of recombination at sites distinct from those that become resolved as crossovers (CO) [24]. We therefore assessed the distribution of RPA, the ssDNA binding protein, which is normally present at focal sites of synapsing meiotic chromosomes before disappearing in mid-pachynema [25]. It is thought to bind D-loops of recombination intermediates [26]. RPA also persisted on pachytene mutant chromosomes (Figure 4K and 4L). These data indicate that unrepaired DSBs, or unresolved recombination intermediates, remain in pachynema and activate a DNA damage checkpoint system."}

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{"project":"2_test","denotations":[{"id":"17696610-16530042-85497902","span":{"begin":707,"end":709},"obj":"16530042"},{"id":"17696610-17446169-85497903","span":{"begin":710,"end":712},"obj":"17446169"},{"id":"17696610-14718568-85497904","span":{"begin":785,"end":787},"obj":"14718568"},{"id":"17696610-16055729-85497905","span":{"begin":788,"end":790},"obj":"16055729"},{"id":"17696610-11950880-85497906","span":{"begin":944,"end":946},"obj":"11950880"},{"id":"17696610-9398850-85497907","span":{"begin":1135,"end":1137},"obj":"9398850"},{"id":"T53052","span":{"begin":707,"end":709},"obj":"16530042"},{"id":"T18220","span":{"begin":710,"end":712},"obj":"17446169"},{"id":"T84823","span":{"begin":785,"end":787},"obj":"14718568"},{"id":"T48623","span":{"begin":788,"end":790},"obj":"16055729"},{"id":"T76001","span":{"begin":944,"end":946},"obj":"11950880"},{"id":"T22662","span":{"begin":1135,"end":1137},"obj":"9398850"}],"text":"Defects in DSB repair became apparent in pachynema upon probing of mutant spermatocyte nuclei with antibodies against molecules involved in various stages of recombination. In \u003e99% of Trip13Gt/Gt chromosome spreads, BLM helicase (Figure 4C and 4D), RAD51/DMC1 (Figure 4E and 4F), γH2AX (Figure 4G and 4H), and TOPBP1 (Figure 4I and 4J) all persisted abnormally on synapsed chromosomes. For RAD51/DMC1, mutant pachytene spermatocytes contained 138 ± 6 foci (compared to 11 ± 3 foci in wild type, most of which were on the XY body), down from 218 ± 13 in zygonema (compared to 220 ± 13 foci in wild type). TOPBP1 is a DNA damage–checkpoint protein involved in ATM protein–dependent activation of ATR protein [20,21]. It binds sites of DSBs and unsynapsed regions of meiotic chromosomes [22,23]. BLM has been reported to colocalize with markers (RPA and MSH4) of recombination at sites distinct from those that become resolved as crossovers (CO) [24]. We therefore assessed the distribution of RPA, the ssDNA binding protein, which is normally present at focal sites of synapsing meiotic chromosomes before disappearing in mid-pachynema [25]. It is thought to bind D-loops of recombination intermediates [26]. RPA also persisted on pachytene mutant chromosomes (Figure 4K and 4L). These data indicate that unrepaired DSBs, or unresolved recombination intermediates, remain in pachynema and activate a DNA damage checkpoint system."}

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in DSB repair became apparent in pachynema upon probing of mutant spermatocyte nuclei with antibodies against molecules involved in various stages of recombination. In \u003e99% of Trip13Gt/Gt chromosome spreads, BLM helicase (Figure 4C and 4D), RAD51/DMC1 (Figure 4E and 4F), γH2AX (Figure 4G and 4H), and TOPBP1 (Figure 4I and 4J) all persisted abnormally on synapsed chromosomes. For RAD51/DMC1, mutant pachytene spermatocytes contained 138 ± 6 foci (compared to 11 ± 3 foci in wild type, most of which were on the XY body), down from 218 ± 13 in zygonema (compared to 220 ± 13 foci in wild type). TOPBP1 is a DNA damage–checkpoint protein involved in ATM protein–dependent activation of ATR protein [20,21]. It binds sites of DSBs and unsynapsed regions of meiotic chromosomes [22,23]. BLM has been reported to colocalize with markers (RPA and MSH4) of recombination at sites distinct from those that become resolved as crossovers (CO) [24]. We therefore assessed the distribution of RPA, the ssDNA binding protein, which is normally present at focal sites of synapsing meiotic chromosomes before disappearing in mid-pachynema [25]. It is thought to bind D-loops of recombination intermediates [26]. RPA also persisted on pachytene mutant chromosomes (Figure 4K and 4L). These data indicate that unrepaired DSBs, or unresolved recombination intermediates, remain in pachynema and activate a DNA damage checkpoint system."}