ArchR Error in (function (..., threads = 1, preschedule = FALSE) : Error Found Iteration 1 :

Thank you very much for developing this tool. However, I encountered a problem while actually running the addReproduciblePeakSet process."

neuron_1 <- addReproduciblePeakSet(
ArchRProj = neuron_1, groupBy = "Clusters", peaksPerCell = 500, pathToMacs2 = pathToMacs2, force = TRUE)

ArchR logging to : ArchRLogs/ArchR-addReproduciblePeakSet-3a341dbd6075-Date-2024-05-13_Time-21-19-56.889968.log If there is an issue, please report to github with logFile!

Calling Peaks with Macs2

2024-05-13 21:19:59.1349 : Peak Calling Parameters!, 0.037 mins elapsed.

Group nCells nCellsUsed nReplicates nMin nMax maxPeaks

C1 C1 318 318 4 63 95 150000 C2 C2 1527 1527 4 352 404 150000 C3 C3 61 55 2 34 37 27500 C4 C4 850 850 4 175 264 150000 C5 C5 472 472 4 88 168 150000 C6 C6 790 790 4 151 229 150000 C7 C7 93 81 2 50 50 40500 C8 C8 522 522 4 98 168 150000 C9 C9 543 543 4 113 151 150000 C10 C10 675 675 4 154 182 150000 C11 C11 730 730 4 166 212 150000 C12 C12 846 846 4 202 220 150000 C13 C13 1443 1443 4 327 386 150000 C14 C14 1519 1519 4 269 485 150000 C15 C15 1444 1444 4 328 387 150000 C16 C16 520 520 4 102 154 150000 C17 C17 556 556 4 113 168 150000 C18 C18 735 735 4 164 219 150000 C19 C19 340 340 4 67 98 150000 C20 C20 322 279 3 59 110 139500 C21 C21 1307 1280 4 134 500 150000 C22 C22 247 247 3 53 106 123500 C23 C23 673 673 4 126 201 150000 C24 C24 516 516 4 114 139 150000 C25 C25 649 649 4 137 183 150000 2024-05-13 21:19:59.148482 : Batching Peak Calls!, 0.038 mins elapsed.

2024-05-13 21:19:59.158763 : Batch Execution w/ safelapply!, 0 mins elapsed.

Warning message in mclapply(..., mc.cores = threads, mc.preschedule = preschedule): “94 function calls resulted in an error” Error in (function (..., threads = 1, preschedule = FALSE) : Error Found Iteration 1 : [1] "Error in data.table::fread(summitsFile, select = c(1, 2, 3, 5)) : \n File '/data/chenwenhai/ATAC5/ATAC_out/PeakCalls/InsertionBeds/C1..WT_M_1101006-1_summits.bed' does not exist or is non-readable. getwd()=='/data/chenwenhai/ATAC5'\n" <simpleError in data.table::fread(summitsFile, select = c(1, 2, 3, 5)): File '/data/chenwenhai/ATAC5/ATAC_out/PeakCalls/InsertionBeds/C1..WT_M_1101006-1_summits.bed' does not exist or is non-readable. getwd()=='/data/chenwenhai/ATAC5'> Error Found Iteration 2 : [1] "Error in data.table::fread(summitsFile, select = c(1, 2, 3, 5)) : \n File '/data/chenwenhai/ATAC5/ATAC_out/PeakCalls/InsertionBeds/C1..CKO_M_1018004-2_summits.bed' does not exist or is non-readable. getwd()=='/data/chenwenhai/ATAC5'\n" <simpleError in data.table::fread(summitsFile, select = c(1, 2, 3, 5)): File '/data/chenwenhai/ATAC5/ATAC_out/PeakCalls/InsertionBeds/C1..CKO_M_1018004-2_summits.bed' does not exist or is non-readable. getwd()=='/data/chenwenhai/ATAC5'> ......

I would greatly appreciate it if you could provide some suggestions.

here is my packagesVersion

[1] SeuratObject_4.1.4 cacoa_0.4.0
[3] Matrix_1.6-3 sp_2.1-2
[5] ArchRtoSignac_1.0.4 ggsci_3.0.3
[7] ggpubr_0.6.0 hexbin_1.28.3
[9] circlize_0.4.16 ComplexHeatmap_2.15.4
[11] dittoSeq_1.17.0 ggrepel_0.9.5
[13] dplyr_1.1.4 ggrastr_1.0.2
[15] harmony_1.2.0 scuttle_1.12.0
[17] SingleCellExperiment_1.24.0 ggridges_0.5.6
[19] nabor_0.5.0 BSgenome.Mmusculus.UCSC.mm10_1.4.3 [21] BSgenome_1.70.1 rtracklayer_1.62.0
[23] BiocIO_1.12.0 Biostrings_2.70.1
[25] XVector_0.42.0 rhdf5_2.46.1
[27] SummarizedExperiment_1.32.0 Biobase_2.62.0
[29] MatrixGenerics_1.14.0 Rcpp_1.0.12
[31] GenomicRanges_1.54.1 GenomeInfoDb_1.38.1
[33] IRanges_2.36.0 S4Vectors_0.40.2
[35] BiocGenerics_0.48.1 matrixStats_1.2.0
[37] data.table_1.14.10 stringr_1.5.1
[39] plyr_1.8.9 magrittr_2.0.3
[41] ggplot2_3.4.4 gtable_0.3.4
[43] gtools_3.9.5 gridExtra_2.3
[45] ArchR_1.0.2

May 13 '24 13:05 wenhai-cell87

Hi @wenhai-cell87! Thanks for using ArchR! Lately, it has been very challenging for me to keep up with maintenance of this package and all of my other responsibilities as a PI. I have not been responding to issue posts and I have not been pushing updates to the software. We are actively searching to hire a computational biologist to continue to develop and maintain ArchR and related tools. If you know someone who might be a good fit, please let us know! In the meantime, your issue will likely go without a reply. Most issues with ArchR right not relate to compatibility. Try reverting to R 4.1 and Bioconductor 3.15. Newer versions of Seurat and Matrix also are causing issues. Sorry for not being able to provide active support for this package at this time.

May 13 '24 13:05 rcorces

Hi @wenhai-cell87,

We encountered a similar issue as yours with summit files not found. In our case, there's no MACS2 files in the output directory at all (ReplicateCalls). This appeared to be MACS2 not running but ArchR did not notice that MACS2 failed and moved on to run the next step.

If this is the same case as ours, running the macs2 callpeaks command in the log file should fail. It worked out after we made sure MACS2 is functional.

Jul 19 '24 17:07 chenyenchung

您好，

我们遇到了与您的问题类似的问题，即找不到 summit files。在我们的例子中，output directory （ReplicateCalls）中根本没有 MACS2 文件。这似乎是 MACS2 没有运行，但 ArchR 没有注意到 MACS2 失败并继续运行下一步。

如果这与我们的情况相同，则在日志文件中运行命令应该会失败。在我们确保 MACS2 正常运行后，它就成功了。macs2 callpeaks

Hello, I've encountered a similar problem. The summit files were not generated correctly. How can I ensure their proper generation? With the same parameters in a certain project, sometimes it works and sometimes it doesn't. So I'm wondering if you have any good ways to ensure that MACS2 generates summit files correctly.

Feb 16 '25 10:02 Superxb1

Hello, I've encountered a similar problem. The summit files were not generated correctly. How can I ensure their proper generation? With the same parameters in a certain project, sometimes it works and sometimes it doesn't. So I'm wondering if you have any good ways to ensure that MACS2 generates summit files correctly.

In our case, MACS2 (or 3) fails due to two main reasons:

MACS2/3 are not in the $PATH ArchR uses. In this case, you would see nothing from MACS.
When running in parallel (multithreading defined by addArchRThreads()), individual processes could be terminated by your system if there is not enough memory. We encountered this as well when running on an HPC without requesting enough resource. This usually manifests as some MACS output directories are fine, while some are either completely missing or missing some files. If this is the case, you'd want to try running ArchR with a smaller number of threads. If you are running it on an HPC, you'd often also see the job terminated with an out-of-memory error while ArchR did not complain.

Feb 17 '25 19:02 chenyenchung

您好，我遇到了类似的问题。未正确生成 summit 文件。如何确保它们的正确生成？在某个项目中使用相同的参数，有时有效，有时无效。所以我想知道你是否有任何好的方法可以确保 MACS2 正确生成峰会文件。

在我们的例子中，（或 3）失败有两个主要原因：MACS2

MACS2/3 未投入使用。在这种情况下，您将看不到来自 .$PATH``ArchR``MACS

当并行运行时（多线程定义），如果内存不足，您的系统可能会终止单个进程。我们在 HPC 上运行时也遇到了这种情况，而没有请求足够的资源。这通常表现为某些 MACS 输出目录正常，而有些则完全缺失或缺少某些文件。如果是这种情况，您需要尝试使用较少数量的线程运行 ArchR。如果您在 HPC 上运行它，您通常还会看到作业因内存不足错误而终止，而 ArchR 没有抱怨。addArchRThreads()

Thank you for your reply. I reduced this parameter to 1, but it still didn't work. It would report an error at the first file, stating that the summits.bed file was not generated. I'm running this on a large - scale computing node. I've confirmed that there is no problem with my memory. Also, other projects can succeed. Occasionally, I encounter such failures. Previously, when I faced this situation, I imported the peakset from cellranger, but I think this is slightly inaccurate.

Feb 19 '25 05:02 superSXB