KEEFEKTIFAN STRATEGI PEMBELAJARAN BERSAFARI BERMINAT, SANGAT MENGUASAI, FAKTA, RABUK PANCAINDRA, DAN DIKSI DALAM PEMBELAJARAN MENULIS CERPEN

Penelitian ini dilatarbelakangi oleh kelemahan siswa dalam menulis cerpen, sehingga diperlukan strategi pembelajaran yang tepat untuk diterapkan dalam pembelajaran menulis cerpen. Berdasarkan latar belakang tersebut, penulis menyusun rumusan masalah: 1) Bagaimana kemampuan siswa dalam menulis cerpen sebelum dan sesudah menggunakan Strategi Bersafari pada kelas eksperimen; 2) Bagaimana kemampuan siswa dalam menulis cerpen sebelum dan setelah tes tanpa menggunakan Strategi Bersafari pada kelas kontrol; 3)Adakah perbedaan yang signifikan antara kemampuan siswa dalam menulis cerpen dengan menggunakan Strategi Bersafari di kelas eksperimen dan tanpa menggunakan Strategi Bersafari di kelas kontrol. Adapun tujuan dari penelitian ini adalah untuk mendeskripsikan hal-hal yang tercantum dalam rumusan masalah tersebut. Strategi pembelajaran yang digunakan dalam penelitian ini adalah Strategi Bersafari. Metode yang digunakan dalam penelitian ini adalah metode eksperimen semu dengan desain pretest-posttest control group. Teknik pengolahan data dalam penelitian ini berupa pengolahan data kuantitatif yang meliputi uji reliabilitas, normalitas, homogenitas, dan uji hipotesis. Terdapat perbedaan yang signifikan antara kemampuan siswa dalam menulis cerpen di kelas eksperimen dan di kelas kontrol. Hal ini bisa dlihat dari pemerolehan hasil posttest antara kedua kelas. Hasil posttest kelas eskperimen sebesar 73, sementara hasil posttest kelas kontrol sebesar 67. Berdasarkan hasil uji hipotesis dengan menentukan thitung, diperoleh nilai thitung = 5,81 dan ttabel = 2,36, sehingga thitung = 5,81 > ttabel = 2,36. Maka hipotesis H1 diterima, yaitu terdapat perbedaan yang signifikan antara kemampuan siswa dalam menulis cerpen di kelas eksperimen dan di kelas kontrol

Acon inactivation and Parkin-mediated mitophagy act in parallel in <i>pink1</i> mutants.

<p>Analysis of <i>parkin</i> mutants (<i>pakin^1/Δ21</i>) and parkin flies heterozygous for <i>acon^1or2</i> (A) Flight ability of 5-day-old adult flies and (B) ATP content in the head-thorax of 5-day-old flies. Data collected from at least 5 independent experiments. (C) GFP-labeled mitochondria in flight muscles (<i>daGal4 UAS-mitoGFP</i>). Scale bar: muscle 10 µm. (D) Quantification of average mitochondrial aggregate size. 5 images from n≥6 thoraxes. (E) Flight ability of control flies, <i>pink1^B9</i>, <i>pink1^B9</i> heterozygous for <i>acon^1or2</i> heterozygous for <i>acon^1or2</i> or overexpressing <i>drp1</i> and <i>pink1</i> mutants with a combination of <i>acon^1or2</i> heterozygousity and <i>drp1</i> overexpression. * Significantly different from <i>pink1^B9</i>, One-way ANOVA, post hoc Dunnett p<0.01, ns: not significantly different.</p

Acon[4Fe-4S] cluster induces mitochondrial defect in DA neurons that is not rescued by increased mitophagy.

<p>(A) Quantification of Western blots of fly heads expressing <i>UAS-acon</i> (see text) in DA neurons probed with anti-Acon normalized to tubulin, relative to control. Data from 3 independent experiments. (B) Mitochondrial Aconitase activity relative to the control. Data collected from 5 independent mitochondrial preparations. (C,E) GFP-labeled mitochondria in dopaminergic (DA) neurons DA neurons labeled with anti-tyrosine hydroxylase (magenta). White arrows indicate mitochondrial aggregates. Scale bar: 2.5 µm. (D,F) Quantification of average mitochondrial aggregate size. 5 neurons per brain from n≥10 brains were analyzed. The genotype of control is: <i>w¹¹¹⁸</i>; <i>pleGal4 UAS-mitoGFP</i>/+ ; and of mutants that express wild type or mutant Acon is: <i>w¹¹¹⁸</i>; <i>UAS-acon^*/+</i>; <i>pleGal4 UAS-mitoGFP</i>/+. Significantly different <b>*</b> from control, ° from ple>acon^wt, °° from ple>acon^S677A One-way ANOVA, post hoc Dunnett p<0.01, ns: not significantly different. Data are shown as Mean ± SEM.</p

Oxidative inactivation of Acon in <i>pink1^B9</i> mutants.

<p>(A) Superoxide production measured as fluorescence change of DHE, a superoxide sensitive dye, in isolated mitochondria from control (<i>pink1^RV</i>), <i>pink1^B9</i>, <i>pink1^B9</i> mutant flies heterozygous for <i>acon^{1 or 2}</i> flies as well as in <i>pink1^RV</i> isolated mitochondria incubated with antimycin. Changes normalized to control. Data collected from 5 independent mitochondrial preparations. * Significantly different from control, One-way ANOVA, post hoc Dunnett p<0.01. (B) Relative mitochondrial Aconitase activity in <i>pink1^B9</i> normalized to Acon protein levels. Data collected from 5 independent mitochondrial preparations. * Significantly different from control, Student's t test p<0.01. (C) H₂O₂ and derivates content measured as H₂O₂-sensitive DCHF fluorescence in fly lysates of indicated genotypes, relative to total protein content. Data collected from 5 independent mitochondrial preparations. * Significantly different from control and ° significantly different from <i>pink1^B9</i>, One-way ANOVA, post hoc Dunnett p<0.01 (D) Fe²⁺ levels in isolated mitochondria from flies with indicated genotype. Quencing of Fe²⁺-sensitive RPA and Fe²⁺-insensitive RPAC dye are represented as percent of initial fluorescence. Data collected from 5 independent mitochondrial preparations. * Significantly different from control and ° significantly different from <i>pink1^B9</i>, One-way ANOVA, post hoc Dunnett p<0.01. Data are shown as Mean ± SEM.</p

Partial loss of Acon suppresses <i>pink1^B9</i> phenotypes.

<p>(A–D) Flight ability of 5-day-old adult flies (A, B) and ATP content in the head-thorax of 5-day-old flies (C, D). Data collected from at least 5 independent experiments. * Significantly different from <i>pink1^B9</i>, One-way ANOVA, post hoc Dunnett p<0.01, ns: not significantly different. (E) TEM analysis of thorax. Black arrows indicate swollen mitochondria. Scale bar: ×5000 5 µm; ×2000 2 µm. (F, H) GFP-labeled mitochondria in flight muscles (F; <i>daGal4 UAS-mitoGFP</i>) and in dopaminergic (H; <i>pleGal4 UAS-mitoGFP</i>) neurons that are double-labeled with anti-tyrosine hydroxylase (magenta). White arrows indicate mitochondrial aggregates. Scale bar: muscle 10 µm; DA neurons 2.5 µm. (G, I) Quantification of average mitochondrial aggregate size. 5 images from n≥6 thoraxes and about 5 neurons per brain from n≥10 brains were analyzed. * Significantly different from <i>pink1^B9</i>, One-way ANOVA, post hoc Dunnett p<0.01, ns not significantly different. In all panel “control” is <i>pink1^RV</i>, a precise <i>P element</i> excision; “genomic” indicates the insertion of a construct on the third chromosome that encompasses the wild type <i>acon</i> gene, data are shown as Mean ± SEM.</p

Model of oxidative Acon inactivation in <i>pink1</i> mutants.

<p>Pink1 loss induces increased superoxide production that inactivates the Acon[4Fe-4S] resulting in the generation of Fe²⁺ and H₂O₂. These combine to form hydroxyl radicals that lead to mitochondrial failure. Mitoferritin chelates Fe²⁺ and is thus able to rescue mitochondrial failure in <i>pink1</i> mutants. Acon inactivation act in parallel to Parkin-mediated mitophagy in controlling mitochondrial integrity.</p

Ndi1p does not rescue <i>parkin</i> mutants.

<p>(A) Quantification of fertility of <i>w</i>; <i>parkin^RV</i> control male flies of <i>w</i>; <i>park^1/Δ21</i> and of <i>w</i>; <i>UAS-NDI1</i>; <i>park^1/Δ21</i> male flies. “RV”: wild type <i>parkin</i> (precise <i>P</i> element excision); “1/Δ21”; mutant <i>park^1/Δ21</i> heteroallelic combination (n = 30 animals). (B) Quantification of flight in control (<i>w</i>; <i>da-Gal4</i>, <i>park^RV</i>) in <i>park^1/Δ21</i> mutant (<i>w</i>; <i>da-Gal4</i>, <i>park^Δ21</i>/<i>park¹</i>) and in <i>park^1/Δ21</i> mutant flies that express Ndi1p (<i>w</i>; <i>UAS-NDI1</i>/<i>+</i>; <i>da-Gal4</i>, <i>park^Δ21</i>/<i>park¹</i>). Student's t-test: * <i>p</i><0.05. Error bars SEM, n = 10 experiments with 5 flies each. (C) GFP labeling of mitochondria from the adult indirect flight muscles of the following genotypes <i>UAS-mito:GFP</i>/<i>+</i>; <i>da-Gal4</i>, <i>park^RV</i> and <i>UAS-mito:GFP</i>/<i>+</i>; <i>da-Gal4</i>, <i>park^Δ21</i>/<i>park¹</i> and <i>UAS-mito:GFP/UAS-NDI1</i>; <i>da-Gal4</i>, <i>park^Δ21</i>/<i>park¹</i>. (D) Quantification of thorax indentations of genotypes indicated in (A). Student's t-test: * <i>p</i><0.03. Error bars SEM, n = 10 experiments with 5 flies each. (E) Complex I enzymatic activity measurement of whole fly-mitochondrial homogenates from <i>pink1^RV</i> controls and <i>pink1^B9</i> mutants compared to <i>park^RV</i> controls and <i>park^1/Δ21</i> mutants. Student's t-test: * <i>p</i><0.05; ns, not significant. Error bars SEM, n>6 experiments with 50 flies each.</p

Increased mitochondrial fission does not rescue reduced Complex I enzymatic activity of <i>pink1</i> mutants.

<p>(A) Quantification of flight in control (<i>w pink1 ^RV</i>) in <i>pink1^B9</i> (<i>w pink1^B9</i>) and in <i>pink1^B9</i> mutant flies that either have a genomic rescue construct of <i>drp1</i> (<i>drp1⁺</i>) or have one mutant allele of <i>opa1</i> (<i>opa1^S3</i>) (<i>w pink1^B9</i>; <i>drp1⁺</i>/+ and <i>w pink1^B9</i>; <i>opa1^S3</i>/+). Student's t-test: ** <i>p</i><0.01; * <i>p</i><0.05. Error bars SEM, n = 10 experiments with 5 flies each. (B) Quantification of thorax indentations of genotypes indicated in (A). Student's t-test: ** <i>p</i><0.01; *** <i>p</i><0.001. Error bars SEM, n = 10 experiments with 5 flies each. (C) Quantification of fertility of <i>w pink1^RV</i> control male flies, <i>w pink1^B9</i> mutants and of <i>w pink1^B9</i>; <i>drp1⁺</i>/+ and of <i>w pink1^B9</i>; <i>opa1^S3</i>/+ resp. male flies (n = 30 animals). (D) Complex I enzymatic activity measurement of whole fly mitochondrial homogenates from <i>w pink1^RV</i> controls and <i>w pink1^B9</i> mutants compared to <i>w pink1^RV</i>; <i>drp1⁺</i>/+ controls, <i>w pink1^B9</i>; <i>drp1⁺</i>/+ mutants and <i>w pink1^RV</i>; <i>opa1^S3</i>/+ controls, <i>w pink1^B9</i>; <i>opa1^S3</i>/+ mutants. Student's t-test: * <i>p</i><0.05. Error bars SEM, n>3 experiments with 50 flies each. The data for <i>w pink1^RV</i> and <i>w pink1^B9</i> is identical to that shown in <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002456#pgen-1002456-g004" target="_blank">Figure 4E</a>. (E) Model of Pink1 and Parkin function in the regulation of mitochondrial activity. Our work indicates that Pink1 has an important function at the level of Complex I, and these defects in part result in mitochondrial remodeling defects seen in the mutants.</p

<i>AOX</i> does not rescue <i>pink1</i> mutants.

<p>(A) Quantitative RT-PCR using primers to <i>CG3446</i> and to <i>AOX</i>, in testes using tissue of the following genotypes <i>w</i>; <i>da-Gal4</i>/<i>+</i> (green), <i>w</i>; <i>UAS-AOX</i> (orange) and <i>w</i>; <i>UAS-AOX</i>/<i>+</i>; <i>da-Gal4</i>/+ (blue). Data normalized to housekeeping genes (see <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002456#pgen.1002456.s006" target="_blank">Text S1</a>). (B) Quantification of fertility of <i>w pink1^RV</i> control male flies of <i>pink1^B9</i>and of <i>w pink1^B9</i>; <i>UAS-AOX</i> male flies (n = 20 animals). (C) Quantification of flight in control (<i>w pink1^RV</i>; <i>da-Gal4</i>/<i>+</i>) in <i>pink1^B9</i> mutant (<i>w pink1^B9</i>; <i>da-Gal4</i>/<i>+</i>) and in <i>pink1^B9</i> mutant flies that express <i>AOX</i> (<i>w pink1^B9</i>; <i>UAS-AOX</i>/+; <i>da-Gal4</i>/<i>+</i>). Student's t-test: ns, not significant. Error bars SEM, n = 6 experiments with 5 flies each. (D) Relative ATP levels of <i>pink1^B9</i> mutant flies (<i>w pink1^B9</i>; <i>da-Gal4/+</i>) normalized to <i>pink1^RV</i> control flies (<i>w pink1^RV</i>; <i>da-Gal4/+</i>) and of <i>pink1^B9</i> mutant flies that express AOX (<i>w pink1^B9</i>; <i>UAS-AOX/+</i>; <i>da-Gal4/+</i>) normalized to control flies that express AOX (<i>w</i>; <i>UAS-AOX/+</i>; <i>da-Gal4/+</i>). Student's t-test: ns, not significant. Error bars SEM, n = 3 experiments with 5 flies each. (E) GFP labeling of mitochondria from the adult indirect flight muscles of the following genotypes <i>w pink1^RV</i>; <i>UAS-mito:GFP</i>/<i>+</i>; <i>da-Gal4</i>/+ and <i>w pink1^B9</i>; <i>UAS-mito:GFP</i>/+; <i>da-Gal4</i>/+ and <i>w pink1^RV</i>; <i>UAS-mito:GFP</i>/<i>UAS-AOX</i>; <i>da-Gal4</i>/+. (F,G) Quantification (F) and labeling (G) of Reserve Pool (RP) vesicles in <i>pink1^RV</i> controls (<i>w pink1^RV</i>; <i>da-Gal4/+</i>) in larvae expressing the AOX transgene (<i>w</i>; <i>UAS-AOX/+</i>; <i>da-Gal4/+</i>), in <i>pink1^B9</i> mutants (<i>w pink1^B9</i>; <i>da-Gal4/+</i>) and in <i>pink1^B9</i> mutants expressing AOX (<i>w pink1^B9</i>; <i>UAS-AOX/+</i>; <i>da-Gal4/+</i>), using the stimulation protocol shown on the left. Student's t-test: * <i>p</i><0.05; ns, not significant. Error bars SEM, n>8 synapses from 4 animals. (H,I) Quantification of red to green JC-1 fluorescence (H) and examples (I) at NMJ boutons in larvae of the genotypes indicated in (F–G). Student's t-test: ns, not significant. Error bars SEM, n>16 synapses from 4 animals.</p